bluuebunny/crossref_metadata_2025 huggingface dataset:
Identification and Characterization of Two DNA Polymerase Activities Present in Trypanosoma brucei Mitochondria
Authors: Fuenmayor Jahely , Zhang Jun , Ruyechan William , Williams Noreen
Published: May 2007

We have identified and partially purified two DNA polymerase activities from purified Trypanosoma brucei mitochondrial extracts. The DNA polymerase activity eluted from the single‐stranded DNA agarose column at 0.15 M KCI (polymerase MI) was significantly inhibited by salt concentrations greater than 100 mM, utilized Mg2+ in preference to Mn2+ as a cofactor on deoxyribonucleotide templates with deoxyribose primers, and in the presence of Mn2+ favored a ribonucleotide template with a deoxyribose primer. A 44 kDa peptide in this fraction crossreacted with antisera against the Crithidia fasciculataβ‐like mitochondrial polymerase. In activity gels the catalytic peptide migrated at an apparent molecular weight of 35 kDa. The DNA polymerase activity present in the 0.3 M KCI DNA agarose fraction (polymerase M2) exhibited optimum activity at 120‐180 mM KCI, used both Mg2+ and Mn2+ as cofactors, and used deoxyribonucleotide templates primed with either deoxyribose or ribose oligomers. Activity gel assays indicate that the native catalytic peptide(s) is ˜ 80 kDa in size. The two polymerases showed different sensitivities to several inhibitors: polymerase MI shows similarities to the Crithidia fasciculataβ‐like mitochondrial polymerase while polymerase M2 is a novel, salt‐activated enzyme of higher molecular weight.
Ultrastructural Features of Cystic and Merogonic Stages of Hepatozoon sipedon (Apicomplexa: Adeleorina) in Northern Leopard Frogs (Rana pipiens) and Northern Water Snakes (Nerodia sipedon) from Ontario, Canada
Authors: Smith Todd G. , Desser Sherwin S.
Published: May 2007

The cystic and merogonic stages of the haemogregarine Hepatozoon sipedon, infecting Northern water snakes (Nerodia sipedon sipedon) and Northern leopard frogs (Rana pipiens), respectively, in Ontario, Canada, were investigated by transmission electron microscopy. Cysts, which were observed in the liver of Northern leopard frogs (Rana pipiens) after these anurans ingested mosquitoes (Culex pipiens) containing oocysts of the parasite, harboured two cystozoites, each of which contained a large crystalloid inclusion anterior to the nucleus. Two types of meronts were observed in snakes that were fed the liver of infected frogs. Macromeronts, which matured in endothelial cells of the liver approximately 16 d after snakes ingested infected frogs, contained about 50 large macromerozoites. Macromerozoites emerged from macromeronts, entered the bloodstream of the snake, and reinfected endothelial cells. Micromeronts, which matured about 34 d post‐inoculation, contained about 150 micromerozoites that infected erythrocytes and transformed into gamonts. The ultrastructural features of micromeronts and macromeronts differed only slightly: immature macromeronts and macromerozoites contained numerous amylopectin and lipid inclusions, whereas immature micromeronts and micromerozoites did not contain amylopectin inclusions and featured fewer, smaller lipid inclusions. A comparison of cystic stages among Hepatozoon species in different groups of vertebrates is presented with respect to their structure and evolutionary significance.
A Comparison of Internal Eliminated Sequences in the Genes that Encode Two K+‐Channel Isoforms in Paramecium tetraurelia
Authors: Ling Kit‐Yin , Vaillant Brian , Haynes W. John , Saimi Yoshiro , Kung Ching
Published: May 2007

We examined both the somatic (macro‐) and the germinal (micronuclear) DNAs that encode two K+‐channel isoforms. PAK1 and PAK11, in Paramecium tetraurelia. The coding regions of these two isoforms are 88% identical in nucleotides and 95% identical in amino acids. Their introns are also highly conserved. Even some of the internal eliminated sequences in PAK1 and PAK11 are clearly related. PAK1 has five IESs; PAK11 has four. The first (5′‐most) IESs of the two genes are located at the same site in the coding sequence but differ in size. The 2nd IES in PAK1 (206‐bp), the largest among the nine IESs, has no PAK11 counterpart. The 3rd, 4th and 5th IESs in PAK1 have a counterpart in PAK11 that is similar in size and in sequence, and identical in its position in the coding sequence. In addition, the first IES of PAK11 bears some resemblance to the 4th one of PAK1. The similarities and differences between the two sets of IESs are discussed with respect to the origin and divergence of the two K+‐channel isoforms.
Molecular Cloning and Characterization of a Superoxide Dismutase (sod) Gene in Pneumocystis carinii
Authors: Denis Cecile‐Marie , Guyot Karine , Wakefield Ann E. , Dive Daniel , Dei‐Cas Eduardo , Camus Daniel , Odberg‐Ferragut Carmen
Published: May 2007

This work reports the isolation and characterization of a gene encoding a superoxide dismutase (SOD. EC.1.15.1.1.) from Pneumocystis carinii derived from rat. Sense and antisense oligonucleotides, deduced from SOD amino acid sequences from a wide variety of organisms, allowed amplification of a 669 bp genomic DNA fragment specific to this P. carinii. RACE‐PCR was used to obtain the major pan of the complementary DNA; the 5‐ and 3′‐genomic regions were obtained respectively from a Mbo I subgenomic library and from an amplified fragment using oligonucleotides designed from the cDNA sequence. Comparison of genomic and cDNA sequences showed an open reading frame of 660 bp interrupted by seven small introns. The deduced amino acid sequence contained 220 residues. Protein sequence alignment demonstrated the highest homology (50.5% identity. 70.3% similarity) with Saccharomyces cerevisiae manganese‐SOD (MnSOD) suggesting that P. carinii SOD belongs to the mitochondrial MnSOD group. A putative targeting peptide found at the 5′‐end of the P. carinii SOD sequence also suggested its mitochondrial localization.
Paramecium tetraurelia Encodes Unconventional Actin Containing Short Introns
Authors: Díaz‐Ramos Corín , Villalobo Eduardo , Pérez‐Romero Pilar , Torres Antonio
Published: May 2007

The polymerase chain reaction was used to amplify and clone an actin gene fragment from Paramecium tetraurelia. This DNA fragment was 1,138 bp long, more than 96% of the actin coding sequence, and contained four in‐frame UAA codons and two small introns located at positions unique in the actin intron catalogue. This is the first report for the phylum Ciliophora of an actin gene containing introns. The deduced amino acid sequence of this actin fragment shared 58‐77% identity with other actins. When compared with rabbit α‐muscle actin, similarities were observed mainly in subdomains 1 and 3, whereas subdomains 2 and 4 appeared to be more divergent.
Structure of the Large Subunit rDNA from a Diatom, and Comparison Between Small and Large Subunit Ribosomal RNA for Studying Stramenopile Evolution
Authors: Auwera Gert Van Der , Wachter Rupert De
Published: May 2007

The aim of this study was to compare the usefulness of complete small and large subunit rRNA, and a combination of both molecules, for reconstructing stramenopile evolution. To this end, phylogenies from species of which both sequences are known Acre constructed with the neighbor‐joining, maximum parsimony, and maximum likelihood methods. Also the use of structural features of the rRNAs was evaluated. The large subunit rRNA from the diatom Skeletonema pseudocostatum was sequenced in order to have a more complete taxon sampling, and a group I intron was identified. Our results indicated that heterokont algae are monophyletic, with diatoms diverging first. However, as the analysis was restricted to a particular data set containing merely six taxa, the outcome has limited value for elucidating stramenopile relationships. On the other hand, this approach permits comparison of the performance of both rRNA molecules without interference from other factors, such as a different species selection for each molecule. For the taxa used, the large subunit rRNA clearly contained more phylogenetic information than the small subunit rRNA. Although this result can definitely not be generalized and depends on the phvlogeny to be studied, in some cases determining complete large subunit rRNA sequences certainly seems worthwhile.
Attachment of Trypanosoma cruzi Epimastigotes to Hydrophobic Substrates and Use of this Property to Separate Stages and Promote Metacyclogenesis
Authors: Kleffmann Torsten , Schmidt Jürgen , Schaub Gunter A.
Published: May 2007

In vivo, epimastigotes of Trypanosoma cruzi colonize a lipidic superficial layer of the rectal cuticle of the vector Triatoma infestans. In vitro, epimastigotes of four cultured strains and one strain from reduviids use a terminal area of the flagellum to attach to a variety of artificial hydrophobic substances, such as hydrocarbons and a range of synthetic plastics. Trypomastigotes did not attach to these substrates. Hydrophilic molecules, such as neutral or negatively charged polysaccharides. did not facilitate binding. Epimastigotes and trypomastigotes were artificially bound by electrostatic forces to positively charged chitosan or DEAE‐Sephacel over their entire surface. Tween 20 and lipid‐binding serum albumin effectively inhibited the hydrophobic attachment. Based on this hydrophobic interaction of epimastigotes. a new chromatography technique has been devised to gently separate trypomastigotes from epimastigotes using octacosane‐coated beads. Furthermore, the in vitro transformation of epimastigotes to trypomastigotes was enhanced if epimastigotes were permitted to attach to hydrophobic, wax‐coated culture vessels.
Description of Two New, Mycophagous Soil Ciliates (Ciliophora, Colpodea): Fungiphrya strobli n. g., n. sp. and Grossglockneria ovata n. sp.
Authors: FOISSNER WILHELM
Published: May 2007

ABSTRACT The morphology and infraciliature of two new, mycophagous soil ciliates are described. Specimens were isolated from dried, rewetted soil samples with the non‐flooded Petri dish method and investigated in vivo and with various silver impregnation techniques. Fungiphrya strobli n. g., n. sp. was discovered in the mud of rock‐pools on the summit of Table Mountain, Republic of South Africa. It is a holotrichously ciliated, about 50 × 40 μm‐sized grossglockneriid with the oral apparatus on the right side of the cell. The adoral ciliature, minute in all other members of the group, is well developed and has a mean of eight kineties forming a conspicuous left, oral polykinetid, highly reminiscent of that found in small species of the genus Colpoda. The ejected extrusomes have a unique, inflated distal end. Grossglockneria ovata n. sp. was discovered in leaf litter from the Lackawanna State Forest in Pennsylvania, USA. It differs from the other members of the genus by the ovate shape, smooth cortex, and the sparse, irregularly‐shaped mucocysts. Taxonomic characters and ranking of grossglockneriids are discussed. Because of the complex, unique feeding tube associated with a unique feeding strategy, mycophagy, it is argued that grossglockneriid ciliates should be classified in a separate order, in spite of their close genetic relatedness to members of the order Colpodida.
Small Subunit Ribosomal RNA Sequence of Henneguya exilis (Class Myxosporea) Identifies the Actinosporean Stage from an Oligochaete Host
Authors: LIN DANJUAN , HANSON LARRY A. , POTE LINDA M.
Published: May 2007

ABSTRACT Several transmission studies, as well as recent molecular data, have indicated that the two classes Myxosporea and Actinosporea represent different life cycle stages of Myxozoa. To evaluate the life cycles of myxozoa in catfish aquaculture systems, the small subunit (18S) ribosomal RNA gene sequences of Henneguya exilis, a myxosporean from channel catfish Ictalurus punctatus, and an actinosporean (previously designated as Aurantiactinomyxon janiszewskai) from the aquatic oligochaete Dero digitata were determined. The sequences were identical, indicating that H. exilis and the actinosporean are alternate life stages of a single species. This is the first report identifying the actinosporean stage of the genus Henneguya.
Encephalitozoon cuniculi: Light and Electron Microscopic Evidence for Di‐, Tetra‐, and Octosporous Sporogony and a Note on the Molecular Phylogeny of Encephalitozoonidae
Authors: VISVESVARA GOVINDA S. , LEITCH GORDON J. , PIENIAZEK NORMAN J.
Published: May 2007

ABSTRACT We demonstrate, based on the light, electron microscopic, and immunofluorescence studies carried out on two isolates of Encephalitozoon cuniculi established in culture, that E. cuniculi exhibits di‐, tri‐, tetra‐ and octosporous sporogony. We therefore propose that the generic characters of Encephalitozoon should be amended to include tetra‐sporous sporogony as generic features. Additionally, the molecular phylogenetic analysis indicates that E. cuniculi, E. hellem, and E. (Septata) intestinalis form a cohesive group.
Rumen Ciliates of White‐tailed Deer (Odocoileus virginianus), Axis Deer (Axis axis), Sika Deer (Cervus nippon) and Fallow Deer (Dama dama) from Texas
Authors: DEHORITY BURK A. , DEMARAIS STEPHEN , OSBORN DAVID A.
Published: May 2007

ABSTRACT Samples of rumen contents from 33 white‐tailed deer (Odocoileus virginianus). 31 axis deer (Axis axis), 26 sika deer (Cervus nippon), and 25 fallow deer (Dama dama) were collected from four study areas in central Texas. The geometric mean concentration of total protozoa was 50.2 times 104 per ml, with no differences between species (P > 0.36). White‐tailed deer had a higher percentage of Entodinium and lower percentage of Diplodiniinae (P≤ 0.01) than the other deer species, which were not different from each other. Occurrence of Epidinium, Isotricha. and Dasytricha was sporadic and did not differ among deer species. Numerous new host records of protozoan species were observed: white‐tailed deer‐four; axis deer‐five; sika deer‐five; fallow deer‐four. This brings the total number of protozoan species identified in each deer species to: white‐tailed‐eight; axis‐12; sika‐15; fallow‐16. For all species combined, protozoan concentrations were 7.5 to 11‐fold higher (P≤ 0.01) from Area 4, which differed from the other three areas by having a stream that allowed deer to have free access to water. Criteria used for identification of medium‐size Eudiplodinium species were evaluated.
Growth, Fine Structure and Cyst Formation of a Microbial Mat Ciliate: Pseudocohnilembus pusillus (Ciliophora, Scuticociliatida)
Authors: OLENDZENSKI LORRAINE C.
Published: May 2007

ABSTRACT Pseudocohnilembus species exhibit a polymorphic life cycle consisting of trophic cells, theronts, and cysts. Pseudocohnilembus pusillus isolated from the intertidal mats of Laguna Figueroa, Baja California Norte, Mexico, forms desiccation‐resistant cysts in response to bacterial food depletion. This isolate is a euryhaline organism, able to grow at salinities from freshwater to 96 ppt total salinity and from pH 6–9. Electron micrographs show that oral and somatic cilia and kinetids are retained inside young cysts. Cyst walls are composed of a single layer (0.1 μm) of granular material. Under all conditions, as bacterial food was depleted, P. pusillus cells formed cysts, except for a small proportion (1–5%) that continued to swim. Changes in pH and salinity did not directly induce cyst formation. Salinity did greatly affect growth rate. Doubling times were shortest at 16 ppt salinity and at pH 7–8. Cyst formation occurred later in the growth cycle as more food bacteria were added. Additionally, ciliates grown in small culture volumes (10 ml) formed cysts sooner than cultures in larger volumes (100 ml), suggesting that crowding may influence cyst formation. Mature cysts may survive desiccation at least as long as one month at 37° C and for as long as one year at 20 ± 3° C. Although trophic cells did not survive desiccation or anoxia, encysted ciliates from liquid stationary phase cultures kept in anoxic seawater for one month excysted into swimming cells within 2.5 h after exposure to air. The adaptability of P. pusillus to extremes of salinity, pH, desiccation, and anoxia permits survival in its environmentally variable, microbial mat habitat.
Abundance of Naked Amoebae in Sediments of Hiroshima Bay, Seto Inland Sea of Japan
Authors: DECAMP OLIVIER , TSUJINO MUTSUMI , KAMIYAMA TAKASHI
Published: May 2007

ABSTRACT The present paper provides the first data on naked amoebae from sediments of Hiroshima Bay. Three stations in the inner part of the bay were sampled over a three‐month period. Abundance of naked amoebae ranged from 1,019 to 45,561 cells/g dry sediment. Results indicate: (i) surface sediment populations in most cases were higher than subsurface populations; (ii) there was some evidence of temporal variation with counts generally increasing from March to May: and (iii) the site located near Hiroshima City had fewer amoebae on several occasions than the other two sites. There was a negative exponential relationship between acid‐volatile sulfide concentration and abundance of amoebae. Most amoebae were small with the average size ranging from 6.6–14 μm. Morphotype 1, amoebae that extend lobose pseudopodia or subpseudopodia during normal locomotion, were dominant (40–100% of enumerated amoebae). Morphotypes 2 and 3 (limax amoebae) were found in lower numbers than the other two morphotypes. The proportion of amoebae occupied by Morphotype 4 (fan‐shaped or discoidal‐flattened amoebae) was higher at a lower total abundance.
Characterization of the Euplotes crassus Macronuclear rDNA and its Potential as a DNA Transformation Vehicle
Authors: ERBEZNIK MILUTIN , YAO MENG‐CHAO , JAHN CAROLYN L.
Published: May 2007

ABSTRACT We have cloned the macronuclear linear DNA molecule carrying the ribosomal RNA genes from the ciliated protozoan Euplotes crassus. DNA sequence analysis was carried out to locate coding regions and to determine whether sequences that have been mutated to confer antibiotic resistance are conserved in the E. crassus genes. The beginning and end of the primary transcript were mapped. In order to determine whether conserved sequences that might serve as replication origins were present, the 5′ and 3′ non‐coding sequences from E. crassus were compared to the corresponding sequences from the macronuclear linear rDNA molecules from the following euplotid species: Euplotes vannus, Euplotes minuta, Euplotes raikovii and Euplotes rariseta. A DNA transformation construct was made by generating a putative anisomycin resistant mutation along with a mutation generating a restriction site polymorphism. Microinjection of the construct into the developing macronucleus of mated cells resulted in exconjugant cell lines with increased resistance to anisomycin. The injected rDNA with the restriction site polymorphism is detectable in the anisomycin resistant cells and appears to represent a minor fraction of the rDNA.
Plastids and Protein Targeting1
Authors: McFADDEN GEOFFREY I.
Published: May 2007

Plastids with two bounding membranes—as exemplified by red algae, green algae, plants, and glaucophytes—derive from primary endosymbiosis: a process involving engulfment and retention of a cyanobacterium by a phagotrophic eukaryote. Plastids with more than two bounding membranes (such as those of euglenoids, dinoflagellates, heterokonts, haptopytes, apicomplexa, cryptomonads, and chlorarachniophytes) probbly arose by secondary endosymbiosis. in which a eukaryotic alga (itself the product of primary endosymbiosis) was engulfed and retained by a phagotroph. Secondary endosymbiosis transfers photosynthetic capacity into heterotrophic lineages, has apparently occurred numerous times, and has created several major eukaryotic lineages comprising upwards of 42.600 species. Plastids acquired by secondary endosymbiosis are sometimes referred to as “second‐hand.” Establishment of secondary endosymbioses has involved transfer of genes from the endosymbiont nucleus to the secondary host nucleus. Limited gene transfer could initially have served to stabilise the endosymbioses, but it is clear that the transfer process has been extensive, leading in many cases to the complete disappearance of the endosymbiont nucleus. One consequence of these gene transfers is that gene products required in the plastid must be targeted into the organelle across multiple membranes: at least three for stromal proteins in euglenoids and dinoflagellates, and across five membranes in the case of thylakoid lumen proteins in plastids with four bounding membranes. Evolution of such targeting mechanisms was obviously a key step in the successful establishment of each different secondary endosymbiosis. Analysis of targeted proteins in the various organisms now suggests that a similar system is used by each group. However, rather than interpreting this similarity as evidence of an homologous origin, I believe that targeting has evolved convergently by combining and recycling existing protein trafficking mechanisms already existing in the endosymbiont and host. Indeed, by analyzing the multiple motifs in targeting sequences of some genes it is possible to infer that they originated in the plastid genome, transferred from there into the primary host nucleus, and subsequently moved into the secondary host nucleus. Thus, each step of the targeting process in “secondhand” plastids recapitulates the gene's previous intracellular transfers.
The Non‐Photosynthetic Plastid in Malarial Parasites and Other Apicomplexans is Derived from Outside the Green Plastid Lineage1
Authors: BLANCHARD JEFFREY L. , HICKS JUSTIN S.
Published: May 2007

The discovery of a non‐photosynthetic plastid genome in Plasmodium falciparum and other apicomplexans has provided a new drug target, but the evolutionary origin of the plastid has been muddled by the lack of characters, that typically define major plastid lineages. To clarify the ancestry of the plastid, we undertook a comprehensive analysis of all genomic characters shared by completely sequenced plastid genomes. Cladistic analysis of the pattern of plastid gene loss and gene rearrangements suggests that the apicomplexan plastid is derived from an ancestor outside of the green plastid lineage. Phylogenetic analysis of primary sequence data (DNA and amino acid characters) produces results that are generally independent of the analytical method, but similar genes (i.e. rpoB and rpoC) give similar topologies. The conflicting phylogenies in primary sequence data sets make it difficult to determine the exact origin of the apicomplexan plastid and the apparent artifactual association of apicomplexan and euglenoid sequences suggests that DNA sequence data may be an inappropriate set of characters to address this phylogenetic question. At present we cannot reject our null hypothesis that the apicomplexan plastid is derived from a shared common ancestor between apicomplexans and dinoflagellates. During the analysis, we noticed that the Plasmodium tRNA‐Met is probably tRNA‐fMet and the tRNA‐fMet is probably tRNA‐Ile. We suggest that P. falciparum has lost the elongator type tRNA‐Met and that similar to metazoan mitochondria there is only one species of methionine tRNA. In P. falciparum, this has been accomplished by recruiting the fMet‐type tRNA to dually function in initiation and elongation. The tRNA‐Ile has an unusual stem‐loop in the variable region. The insertion in this region appears to have occurred after the primary origin of the plastid and further supports the monophyletic ancestory of plastids.
Parasitic Life Styles of Marine Dinoflagellates1
Authors: COATS D. WAYNE
Published: May 2007

Several genera of marine dinoflagellates contain species that have evolved parasitic life styles. Dinoflagellate infections have been reported for a wide range of host organisms including sarcodines. ciliates, free‐living dinoflagellates, various invertebrates, and a few vertebrates. Some dinoflagellates even parasitize other parasitic dinoflagellates. Most species are obligately parasitic and rely on heterotrophy as their sole means of nutrition; however, some are mixotrophic, as they possess chloroplasts during part or all of their life cycle. Many are ectoparasites that use highly specialized structures to attach to their host and feed, while others are intracellular parasites that feed by osmotrophy. Parasitic dinoflagellates often have adverse effects on their host that can lead to reproductive castration or death. The ecological importance of parasitic dinoflagellates is particularly evident during epidemic outbreaks that cause mass mortality of host organisms. Species that infect fish can pose threats to aquaculture. while other species can make commercially important crustacea unpalatable. In the planktonic realm, parasitic dinoflagellates influence the structure and function of the microbial food web. They compete with copepods and other grazers by utilizing ciliates as hosts and can stimulate rapid recycling of nutrients by causing the decline of toxic and non‐toxic red tides.
Voltage‐Dependent Reversal of Anodic Galvanotaxis in Nyctotherus ovalis
Authors: HOEK ANGELA H. A. M. VAN , SPRAKEL VERA S. I. , ALEN THEO A. VAN , THEUVENET ALEXANDER P. R. , VOGELS GODFRIED D. , HACKSTEIN JOHANNES H. P.
Published: May 2007

Aerobic and anaerobic ciliates swim towards the cathode when they are exposed to a constant DC field. Nyctotherus ovalis from the intestinal tract of cockroaches exhibits a different galvanotactic response: at low strength of the DC field the ciliates orient towards the anode whereas DC fields above 2–4 V/cm cause cathodic swimming. This reversal of the galvanotactic response is not due to backward swimming. Rather the ciliates turn around and orient to the cathode with their anterior pole. Exposure to various cations, chelators, and Ca2‐‐channel inhibitors suggests that Ca2‐‐channels similar to the “long lasting” Ca2‐‐channels of vertebrates are involved in the voltage‐dependent anodic galvanotaxis. Evidence is presented that host‐dependent epigenetic factors can influence the voltage‐threshold for the switch from anodic to cathodic swimming.
Fractionation of Sporogonial Stages of the Microsporidian Encephalitozoon cuniculi by Percoll® Gradients
Authors: GREEN LINDA C. , DIDIER PETER J. , DIDIER ELIZABETH S.
Published: May 2007

Microsporidia are obligate intracellular parasites that are increasingly recognized as a cause of opportunistic infections in immunocompromised individuals. Encephalitozoon cuniculi has been identified in humans with AIDS and infects a wide range of mammalian hosts. Little is known about the metabolic processes that regulate growth and replication of microsporidia. Examination of the individual stages of development will facilitate such studies and reveal possible targets for drug therapy. The purpose of this study was to fractionate and purify stages of the microsporidian life cycle. Encephalitozoon cuniculi were cultured in RK‐13 cells. The tissue supernatants containing multiple parasite stages, empty microsporidial husks and host cell debris were collected, washed, and subjected to differential centrifugation in 80% stock isotonic Percoll®. Transmission electron microscopy and SDS‐polyacrylamide gel electrophoresis were used to compare the content and purity of each fraction. Mature spores formed a band at a density of approximately 1.138 g/ml. Sporoblasts were found at densities between 1.102 g/ml and 1.119 g/ml. A mixture of sporonts, sporoblasts, microsporidial husks, and cell debris remained at the top of the gradient and additional centrifugation in 30% and 50% Percoll® resulted in separation of these stages. These results represent the first step toward fractionating stages of microsporidia infecting humans.
Characterization of Two Perkinsus spp. from the Softshell Clam, Mya arenaria Using the Small Subunit Ribosomal RNA Gene
Authors: KOTOB SHABAN I. , McLAUGHLIN SHAWN M. , BERKUM PETER van , FAISAL MOHAMED
Published: May 2007

Sequence analysis and riboprinting of the small subunit ribosomal RNA genes were used to characterize two morphologically different Perkinsus species isolates from the gill (G117) and the hemolymph (H49) of the softshell clam, Mya arenaria. Sequence data of the polymerase chain reaction amplified ribosomal RNA loci of G117 and H49 indicated that these genes are 1803 and 1806 base‐pair long, respectively. A sequence similarity of > 98.9% was calculated among ribosomal RNA sequences of the two isolates of this study and the published sequences of Perkinsus marinus from the American eastern oyster, Crassostrea virginica, and Perkinsus sp. from the blood cockle of the Australian mollusc, Anadara trapezia. From a phylogenetic tree obtained from Jukes‐Cantor distances of the aligned ribosomal RNA gene sequences of 13 eukaryotic taxa using the Neighbor‐Joining method, we showed that G117 and H49 clustered within the genus Perkinsus. Guided by the sequence data of Perkinsus marinus (accession # X75762) and Perkinsus sp. (accession # L07375), restriction endonucleases were selected for restriction fragment analysis of polymerase chain reaction products of the small subunit ribosomal RNA genes (riboprinting). Riboprinting was used to distinguish the four members of the genus Perkinsus from each other.
Keto‐Acid Oxidoreductases in the Anaerobic Protozoa1
Authors: UPCROFT JACQUELINE A. , UPCROFT PETER
Published: May 2007

In anaerobes, decarboxylation of pyruvate is executed by the enzyme pyruvate: ferredoxin oxidoreductase, which donates electrons to ferredoxin. The pyruvate:ferredoxin oxidoreductase and its homologues utilise many alternative substrates in bacterial anaerobes. The pyruvate:ferredoxin oxidoreductase from anaerobic protozoa, such as Giardia duodenalis, Trichomonas vaginalis, and Entamoeba histolytica have retained this diversity in usage of alternative keto acids for energy production utilising a wide variety of substrates. In addition to this flexibility, both T. vaginalis and G. duodenalis have alternative enzymes that are active in metronidazole‐resistant parasites and that do not necessarily involve donation of electrons to characterized ferredoxins. Giardia duodenalis has two oxoacid oxidoreductases, including pyruvate:ferredoxin oxidoreductase and T. vaginalis has at least three. These alternative oxoacid oxidoreductases apparently do not share homology with the characterized pyruvate:ferredoxin oxidoreductase in either organism. Independently, both G. duodenalis and T. vaginalis have retained alternative oxoacid oxidoreductase activities that are clearly important for the survival of these parasitic protists.
Analysis of Genomic G + C Content, Codon Usage, Initiator Codon Context and Translation Termination Sites In Tetrahymena Thermophila
Authors: WUITSCHICK JEFFREY D. , KARRER KATHLEEN M.
Published: May 2007

ABSTRACT. In recent years, the amount of molecular sequencing data from Tetrahymena thermophila has dramatically increased. We analyzed G + C content, codon usage, initiator codon context and stop codon sites in the extremely A + T rich genome of this ciliate. Average G + C content was 38% for protein coding regions. 21% for 5′ non‐coding sequences, 19% for 3′ non‐coding sequences, 15% for introns, 19% for micronuclear limited sequences and 17% for macronuclear retained sequences flanking micronuclear specific regions. the 75 available T. thermophila protein coding sequences favored codons ending in T and, where possible, avoided those with G in the third position. Highly expressed genes were relatively G + C‐rich and exhibited an extremely biased pattern of codon usage while developmentally regulated genes were more A + T‐rich and showed less codon usage bias. Regions immediately preceding Tetrahymena translation initiator codons were generally A‐rich. For the 60 stop codons examined, the frequency of G in the end + 1 site was much higher than expected whereas C never occupied this position.
Correlation Between Loss of A Mg2+ Conductance and an Adaptation Defect In A Mutant of Paramecium Tetraurelia
Authors: PRESTON ROBIN R. , HAMMOND JOCELYN A.
Published: May 2007

ABSTRACT. Paramecium tetraurelia responds to chronic KCI‐induced depolarization by swimming backward, but the ciliate recovers within seconds and then undergoes a prolonged adaptation period during which sensitivity to external stimuli is altered radically. We examined the role of Mg2‐ in this phenomenon, prompted by finding that mutations in the eccentric‐A gene both suppressed a Mg2+‐ specific conductance and prevented adaptation. Adaptation of the wild type proceeded normally when extracellular Mg2‐ was varied from 0‐20 mM, however, suggesting that channel‐mediated Mg2+ fluxes were not involved. In seeking alternative explanations for the eccentric mutant phenotype, we ascertained that there was an osmotic component to adaptation but that K+‐induced depolarization was the primary stimulus. We also noted that wild‐type and eccentric mutant cells depolarized by equivalent amounts in KCI, suggesting that the genetic lesion must lie downstream of membrane‐potential change. We also examined whether the adaptation‐induced behavioral changes and, indeed, the defect in eccentric might be explained in terms of Mg2+ and Na+ efflux during behavioral testing, but experimental observations failed to support this notion. Finally, we consider the possibility that eccentric gene mutation prevents adaptation by interfering with intracellular free Mg2+ homeostasis in Paramecium.
Babesia Canis Canis, Babesia Canis Vogeli, Babesia Canis Rossi: Differentiation of the Three Subspecies By A Restriction Fragment Length Polymorphism Analysis On Amplified Small Subunit Ribosomal Rna Genes
Authors: CARRET CÉLINE , WALAS FABIEN , CARCY BERNARD , GRANDE NATHALIE , PRÉCIGOUT ÉRIC , MOUBRI KARINA , SCHETTERS THEO P. , GORENFLOT ANDRÉ
Published: May 2007

ABSTRACT. The parasites Babesia canis and Babesia gibsoni (phylum Apicomplexa) are responsible for canine babesiosis throughout the world. Babesia canis was previously described as a group of three biologically different subspecies, namely B. canis canis. B. canis vogeli, and B. canis rossi. We report partial sequences of small subunit ribosomal RNA gene (ssu‐rDNA) of each subspecies amplified in vitro with primers derived from a semi‐conserved region of the ssu‐rDNA genes in other Babesia species. the polymerase chain reaction combined with a restriction fragment length polymorphism analysis, using Hinfl and Taql restriction enzymes, confirmed the separation of B. canis into three subspecies. These sequences were compared with previously published sequences of other Babesia species. A phylogenetic approach showed that the three subspecies of B. canis belong to the clade of Babesia species sensu stricto where B. canis canis clusters with B. canis rossi whereas B. canis vogeli might form a monophyletic group with the cluster B. divergens and B. odocoilei. Our results show that the three subspecies of B. canis can readily be differentiated at the molecular level and suggest that they might be considered as true species.
A Cytogenetic Study of Development In Mechanically Disrupted Pairs of Tetrahymena Thermophila
Authors: VIRTUE MARK A. , COLE ERIC S.
Published: May 2007

ABSTRACT We examined the nuclear behavior of mating Tetrahymena cells that had been mechanically disrupted at various times throughout conjugation. Disruption was achieved by agitating conjugating Tetrahymena in the presence of 0.1‐3 mm glass beads. Two minutes of agitation with 1 mm beads yielded optimal pair disruption (70%) with high viability (92%). Disrupting pairs between 0‐4.7 h after the initiation of mating produced mostly disrupted conjugants in which development was aborted. However, as many as 20% of these early disrupted conjugants completed development even without their mating partners. After 5 h the percentage of disrupted conjugants completing development increased dramatically, reaching 80% by 6.7 h. These results support a model suggesting that events associated with nuclear exchange and fusion 5 h into conjugation trigger a commitment to completion of the postzygotic developmental program. the early conjugants that completed development following disruption suggest that development can be sustained even in the absence of a mating partner. This represents a novel method of bringing the micronuclear genome into macronuclear expression with minimal cytoplasmic exchange between partners. We discuss these results in light of a model relating cortical and nuclear signaling events that reciprocally drive conjugal development.
Actin of Histriculus cavicola: Characteristics of the Highly Divergent Hypotrich Ciliate Actins1
Authors: PÉREZ‐ROMERO PILAR , VILLALOBO EDUARDO , DÍAZ‐RAMOS CORÍN , CALVO PURIFICACIÓN , TORRES ANTONIO
Published: May 2007

A macronuclear gene‐sized molecule carrying an actin gene from the hypotrich ciliate, Histriculus cavicola, was characterized. Southern blot analysis using a coding region probe suggested that actin in H. cavicola is encoded by a single gene. A comparison of the promoter regions indicated that the H. cavicola actin gene has a TATA box in the 51 flanking region in a position identical to those in other oxytrich ciliates. The coding sequence of this gene is not interrupted by any introns, and codes for a protein of 375 amino acid residues. This protein shares a high degree of similarity with other oxytrichid actins, and a relatively low similarity with actins from other eukaryotes. Comparative analyses of sequences indicated that most of the amino acid substitutions in hypotrich actins are found in surface loops, while the core structures are well‐conserved. The sites that interact with DNase I and several regions involved in actin‐actin contact have diverged considerably in hypotrich actins, while nucleotide‐binding sites are the best‐conserved interaction motif.
Ultrastructural Characterisation and Molecular Taxonomic Identification of Nosema granulosis n. sp., a Transovarially Transmitted Feminising (TTF) Microsporidium
Authors: TERRY REBECCA S. , SMITH JUDITH E. , BOUCHON DIDIER , RIGAUD THIERRY , DUNCANSON PHIL , SHARPE ROSE G. , DUNN ALISON M.
Published: May 2007

A novel microsporidian parasite is described, which infects the crustacean host Gammarus duebeni. The parasite was transovarially transmitted and feminised host offspring. The life cycle was monomorphic with three stages. Meronts were found in host embryos, juveniles, and in the gonadal tissue of adults. Sporoblasts and spores were restricted to the gonad. Sporogony was disporoblastic giving rise to paired sporoblasts, which then differentiated to form spores. Spores were not found in regular groupings and there was no interfacial envelope. Spores were approximately 3.78 × 1.22 μm and had a thin exospore wall, a short polar filament, and an unusual granular polaroplast. All life cycle stages were diplokaryotic. A region from the parasite small subunit ribosomal RNA gene was amplified and sequenced. Phylogenetic analysis based on these data places the parasite within the genus Nosema. We have named the species Nosema granulosis based on the structure of the polaroplast.
A Novel Healing Filament in Ciliate Regeneration
Authors: JERKA‐DZIADOSZ MARIA , MUSZYNSKA KATARZYNA , KRAWCZYńSKA WANDA
Published: May 2007

The interphase cells of the hypotrich ciliate Paraurostyla weissei possess a complex fibrillar system surrounding basal bodies in the compound ciliary assemblages, cirri and membranelles. During replacement of the ciliature at cell division, transient filaments precede and accompany the development of ciliary primordia and participate in the formation of the fission furrow. Both fibrillar systems are recognized by monoclonal antibody FXXXIX 12G9. We studied regeneration of cellular fragments after transection employing the mAb 12G9 and found a new cytoskeletal structure involved in healing of the excisional wound. The healing filament is formed at the wound edge, distally and in connection with the bases of cirri closest to the wound. It is visible 5 min after transection. Concomitant with development of new ciliary primordia, the healing filament shrinks and finally disappears together with other transient fibers formed in this process. Ultrastructural analysis of immunolabeled regenerating cells revealed that structures recognized by mAbl2G9 contain fine filaments whose packing and arrangement depends on accompanying cytoplasmic elements and the developmental status of a fragment. Assembly of the healing fiber does not depend on microtubules and microfilaments since it develops in cellular fragments exposed to cold, nocodazole, and Cytochalasin D. On Western blots of whole cell and cytoskeletal extracts of P. weissei the 12G9 antibody identified one protein band whose molecular weight corresponds to 60 kDa.
Social Comparison and Satisfaction: Students' Reactions After Exam Feedback Predict Future Academic Performance1
Authors: Lane David J. , Gibbons Frederick X.
Published: May 2007

These studies examined the relations among performance satisfaction, choice of social comparison targets, and subsequent performance in a classroom setting. Previous research has shown that comparison with high performers helps academic performance. In 2 field studies, this relation was moderated by satisfaction with initial performance. After learning their scores on the first exam of the semester, students indicated their satisfaction with the scores and performance level of someone with whom they would like to compare exams. Comparison with poor performers was beneficial when coupled with dissatisfaction. Dissatisfied students who preferred comparison with poor performers improved over the semester.
Smoking interferes with the prognosis of dental implant treatment: a systematic review and meta‐analysis
Authors: Strietzel Frank Peter , Reichart Peter A. , Kale Abhijit , Kulkarni Milind , Wegner Brigitte , Küchler Ingeborg
Published: May 2007

Aim: This systematic literature review was performed to investigate if smoking interferes with the prognosis of implants with and without accompanying augmentation procedures compared with non‐smokers.

Methods: A systematic electronic and handsearch (articles published between 1989 and 2005; English and German language; search terms “dental or oral implants and smoking”; “dental or oral implants and tobacco”) was performed to identify publications providing numbers of failed implants, related to the numbers of smokers and non‐smokers for meta‐analysis. Publications providing statistically examined data of implant failures or biologic complications among smokers compared with non‐smokers were included for systematic review.

Results: Of 139 publications identified, 29 were considered for meta‐analysis and 35 for systematic review. Meta‐analysis revealed a significantly enhanced risk for implant failure among smokers [implant‐related odds ratio (OR) 2.25, confidence interval (CI95%) 1.96–2.59; patient‐related OR 2.64; CI95% 1.70–4.09] compared with non‐smokers, and for smokers receiving implants with accompanying augmentation procedures (OR 3.61; CI95% 2.26–5.77, implant related). The systematic review indicated significantly enhanced risks of biologic complications among smokers. Five studies revealed no significant impact of smoking on prognosis of implants with particle‐blasted, acid‐etched or anodic oxidized surfaces.

Conclusion: Smoking is a significant risk factor for dental implant therapy and augmentation procedures accompanying implantations.
Four‐year results of a prospective‐controlled clinical study evaluating healing of intra‐bony defects following treatment with an enamel matrix protein derivative alone or combined with a bioactive glass
Authors: Sculean Anton , Pietruska Malgorzata , Arweiler Nicole B. , Auschill Thorsten M. , Nemcovsky Carlos
Published: May 2007

Aim: To evaluate the 4‐year results following regenerative periodontal surgery at intra‐bony defects with either a combination of an enamel matrix protein derivative (EMD) and a bioactive glass (BG) or with EMD alone.

Methods: Twenty‐five patients with one deep intra‐bony defect each were randomly treated with either an EMD+BG (test) or with EMD alone (control). Measurements were recorded at baseline, at 1 and at 4 years following therapy. The primary outcome variable was the clinical attachment level (CAL).

Results: The test group demonstrated a mean CAL change from 10.3±1.6 to 6.7±1.2 mm (p<0.001) and to 6.9±1.0 mm (p<0.001) at 1 and 4 years, respectively. No statistically significant differences were found between the 1‐ and 4‐year results. The control group showed a mean CAL change from 10.4±1.6 to 6.7±1.1 mm (p<0.001) at 1 year and 7.0±0.9 mm (p<0.001) at 4 years. The CAL change between 1 and 4 years did not present statistically significant differences. In each of the two groups, four defects have lost 1 mm of the CAL gained at 1 year. A CAL gain of 1 mm compared with the 1‐year results was measured in only one defect of the test group. Compared with baseline, a CAL gain of 3 mm was found at 4 years in 10 defects in both groups.

Between the treatment groups, no statistically significant differences in any of the investigated parameters were observed at 1 and at 4 years.

Conclusions: Within their limits, the present results indicate that the clinical improvements obtained with both regenerative modalities can be maintained over a period of 4 years.
Single‐nucleotide polymorphisms in the IL‐4 and IL‐13 promoter region in aggressive periodontitis
Authors: Gonzales J. R. , Mann M. , Stelzig J. , Bödeker R. H. , Meyle J.
Published: May 2007

Introduction: IL‐4 and IL‐13 polymorphisms have been shown to influence the susceptibility to systemic diseases. In this study, possible associations between the IL‐4 −590 C→T, IL‐4 −34 C→T, IL‐13 −1112 C→T and IL‐13 −1512 A→C promoter polymorphisms were investigated in subjects with generalized aggressive periodontitis (AgP) compared with healthy individuals.

Material and Methods: Fifty‐eight patients with diagnosis of generalized AgP and 51 matched healthy controls participated in the study. Blood samples were collected and DNA isolated. Molecular analyses were performed by PCR‐RFLP in a blind fashion. Genotype and allele frequencies among study groups were compared using Fisher's exact test (α value: 0.05). Pearson's χ2 test was used for analysis of Hardy–Weinberg equilibrium.

Results: The frequency of the IL‐4 −590 T/T and IL‐4 −34 T/T genotypes differed significantly between groups (p=0.05, 0.02, respectively), although the allele frequencies were similar. There was a higher frequency of the IL‐4 −590 T/T and IL‐4 −34 T/T genotypes in patients with AgP compared with controls. The genotype and allele frequencies of the IL‐13 polymorphisms did not differ between groups.

Conclusions: This study demonstrated an association between the IL‐4 −590 T/T and IL‐4 −34 T/T genotypes and AgP. Further research is necessary to prove if there is an association of these polymorphisms with AgP, and if the polymorphisms have a functional effect.