bluuebunny/crossref_metadata_2025 huggingface dataset:
Ultrastructure of Prorodon (Ciliophora, Prostomatida) Ii. Oral Cortex and Phylogenetic Conclusions
Authors: HILLER STEPHAN A.
Published: May 2007

ABSTRACT. The ultrastructure of the oral region and the ultrastructural architecture of the basket of Prorodon aklitolophon and Prorodon teres are described. the oral region of Prorodon consists of: 1) A circumoral kinety at the anterior pole of the cell surrounding the typically slit‐shaped cytostomial funnel. This kinety is composed of inversely oriented dikinetids in which both kinetosomes are ciliated and are associated with a very short postciliary microtubular ribbon and a few transverse microtubules; 2) Three brush rows aligned in parallel and extended meridionally in the anterior part of the cell. the individual brush rows consist of dikinetids, but in contrast to the dikinetids around the cytopharynx they are not inverted and only the anterior kinetosomes bear specialized short brush cilia and are associated with a divergent‐tangential transverse microtubular ribbon. the posterior kinetosome is non‐ciliated and bears a prominent convergent postciliary microtubular ribbon. Schematized dikinetid patterns of both oral regions of Prorodon are provided. In addition, a three‐dimensional reconstruction of the basket of the genus Prorodon based on serial thin sections is presented. A phylogenetic tree, mainly based on stomatogenic data, is given to show the phylogenetic relationships of some prostomatid genera as well as the hypothesized sistergroup relationship of colpodid and prostomatid ciliates.
The Dynamic Nature of Protistan Ingestion Response to Prey Abundance
Authors: CHOI JOON W.
Published: May 2007

ABSTRACT. Ingestion rate of Paraphysomonas imperforata was found to be a hyperbolic function of prey density. But the same flagellate clone had multiple ingestion responses to prey density, depending on its physiological state and physical stress it suffers. The flagellates in a physiological state of higher growth tended to have higher maximum ingestion and clearance rates than ones in a physiological state of lower growth. The same trend was observed for volume‐specific maximum ingestion and volume‐specific clearance rate. In response to changing prey density, the growth rate did not change as quickly as the ingestion rate, suggesting imbalance between the two. The tested physical stresses, including shaking, centrifugation, and filtration, also resulted in reduction of ingestion parameters of the flagellates. But half‐saturation constants did not show any trend in response to either physiological state or physical stress. In light of the dynamic nature of protistan ingestion response to prey abundance, short incubation, which minimizes the physiological change, and careful handling, which prevents the possible physical stress, should be employed in order to avoid underestimation of in situ ingestion rates. Previously reported ingestion parameters of lab‐cultured protists, which are thought to be unrealistic in natural conditions, may represent only one of multiple ingestion responses, probably prey‐rich condition.
On Paraptychostomum almae N. G., N. Sp., a Commensal Ciliate from the Digestive Tract of Oligochaetes of the Cameroons, in a New Subclass Hysterocinetia
Authors: NGASSAM PIERRE , DE PUYTORAC PIERRE , GRAIN JEAN
Published: May 2007

ABSTRACT. Morphological and ultrastructural studies on a new ciliate, Paraptychostomum almae, from the digestive tract of an oligochaete (Alma emini) from the Cameroons are carried out. The flattened cell has a large size; its left lateral face bears an anterior thigmotactic zone that includes seven‐nine short kinetal segments. The somatic cortex is composed of flattened alveoli, a thin epiplasm and a microfibrillar ecto‐endoplasmic boundary. Kineties are made of monokinetids, each particularly characterized by a long anteriorly directed kinetodesmal fiber, and a hyperdivergent postciliary ribbon. The postero‐ventral buccal apparatus consists of a short peristome and a deep longitudinal infundibulum. The paroral organelle is a long stichodyad. The three adoral organelles are of different types: ADI and AD3 are of the membranoid type, respectively with two and one rows of ciliated kinetosomes; AD2 is of the peniculus type with six‐seven rows of ciliated kinetosomes. A microfibrillar network with nodes arises from all the buccal kinetosomes and extends under the naked wall. Mitochondria are small and numerous and dispersed throughout the whole cell. The existence of an AD2 with more than two rows of kinetosomes warrants the creation of the new genus Paraptychostomum and a new family, Ptychostomatidae. The presence of a distinct ecto‐endoplasmic boundary and of somatic kinetids exclusive without transversal dense tractus, hyperdivergent postciliary ribbons, and dispersed numerous mitochondria, added to particularities of the stomatogenesis, allow us to clearly separate hysterocinetians from the scuticociliates and to set up for them the new subclass Hysterocinetia, within the class Oligohymenophorea, with a single new order Hysterocinetida.
Investigations on the Host Specificity of Dihomoxenous Sarcosporidia in the Intermediate and Definitive Host
Authors: BANNERT BRIGITTE
Published: May 2007

ABSTRACT. Cross‐transmission experiments were performed in order to determine the host specificity in the intermediate and definitive hosts of the four described dihomoxenous Sarcocystis species, S. gallotiae, S. stehlinii, S. simonyi, and S. dugesii from lacertid lizards of the genera Gallotia and Podarcis from the Macaronesian Islands. Sarcocysts of either species from experimentally infected lizards were fed to a variety of laboratory‐bred lizard species of the genera Gallotia, Lacerta, and Podarcis. These sarcocysts proved to be infectious to all examined animals, showing no definitive host specificity in the tested genera. Lizards of the genera Chalcides and Tarentola, however, were not susceptible definitive hosts for S. gallotiae. The inoculation of experimentally obtained sporocysts of each of the four Sarcocystis species to various lacertid lizard species revealed varying degrees of intermediate host specificity, generally demonstrating each native host to be the most susceptible.
Expression of Cysteine Proteinases by Metacyclic Promastigotes of Leishmania mexicana
Authors: BATES PAUL A. , ROBERTSON COLIN D. , COOMBS GRAHAM H.
Published: May 2007

ABSTRACT. The expression of cysteine proteinases by metacyclic promastigotes of Leishmania mexicana was investigated using gelatin polyacrylamide gel electrophoresis. Two prominent bands were detected which distinguished metacyclics from multiplicative promastigotes, lacking detectable cysteine proteinase activity, and amastigotes, with a distinct banding pattern composed of multiple enzymes. A correlation between relative activity of the metacyclic‐specific bands and the prevalence of metacyclics was found both during the growth cycle in vitro as metacyclogenesis occurred, and by comparison of stationary phase populations from consecutive subpassages in vitro. Irreversible inhibition of the metacyclic activities using N‐benzyloxycarbonyl‐phenylalanyl‐alanyl diazomethane did not inhibit metacyclic to amastigote transformation in vitro. These activities provide a useful biochemical marker for the metacyclic promastigotes of L. mexicana.
Comparative Study of Microsporidian Spores by Flow Cytometric Analysis
Authors: AMIGÓ JOSEP M. , GRACIA MARIA‐PILAR , COMAS JAUME , SALVADÓ HUMBERT , VIVARÈS CHRISTIAN P.
Published: May 2007

ABSTRACT. Spore suspensions of microsporidian parasites of fish (Microsporidium ovoideum, Glugea stephani, Glugea atherinae and Spraguea lophii) have been analyzed by flow cytometry. Spore nuclei were dyed either by propidium iodide or bis‐benzimide (Hoechst 33342). By observation of forward light scatter and fluorescence the four species could be distinguished and the mono‐ and diplokaryotic populations of S. lophii identified. Staining of DNA by bis‐benzimide was better and easier than propidium iodide. Forward light scatter and fluorescence values were characteristic of each species and remained unchanged throughout the year, so flow cytometry can be used for distinction of spores of some microsporidian parasites once their flow cytometric parameters are known. However, special care has to be taken in tool calibration and material preparation for analysis because of the high precision of the technique.
Secretion of Sucrase by Leishmania donovani
Authors: BLUM J. JOSEPH , OPPERDOES FRED R.
Published: May 2007

ABSTRACT. Leishmania donovani promastigotes were collected, washed, resuspended in buffer, and assayed for sucrase activity. No activity was observed in the intact washed cells, but activity was measurable when the cells were permeabilized with Triton X‐100. Intracellular sucrase activity was highest in promastigotes grown at pH 7.4, somewhat lower in promastigotes grown at pH 5.5, and significantly lower in “amastigotes” grown at pH 5.5. No trehalase, lactase, or maltase activities were observed. Assay of the medium in which the cells had grown showed that most the sucrase activity was extracellular, i.e. was secreted into the medium during growth.
Randomly Amplified Polymorphic DNA (RAPD) and Isoenzyme Analysis of Trypanosoma rangeli Strains
Authors: STEINDEL MARIO , NETO EMMANUEL DIAS , PINTO CARLOS J. CARVALHO , GRISARD EDMUNDO C. , MENEZES CARLA L. P. , MURTA SILVANE M. F. , SIMPSON ANDREW J. G. , ROMANHA ALVARO J.
Published: May 2007

ABSTRACT. Sixteen Trypanosoma rangeli strains were compared by isoenzyme and randomly amplified polymorphic DNA (RAPD) analysis. Eight strains were isolated from either Rhodnius prolixus or Homo sapiens from Honduras, Colombia and Venezuela. Another eight strains were isolated from either Panstrongylus megistus or the rodent Echimys dasythrix from the State of Santa Catarina, southern Brazil. All six T. rangeli strains isolated from P. megistus were co‐infections with Trypanosoma cruzi, demonstrating an overlap of the sylvatic cycles of these parasites and that the accurate identification of species is of utmost importance. Both isoenzyme and RAPD analysis revealed two distinct groups of T. rangeli strains, one formed by the strains from Santa Catarina and the other, by the strains from Honduras, Colombia and Venezuela. With the five enzymes used, all the strains from Santa Catarina had identical profiles which overlapped with those of the other regions only in the pattern obtained with malic enzyme. Analysis of 138 RAPD bands by means of an unweighted pair group method analysis (UPGMA) phenogram using the Dice similarity coefficient allowed the separation of the two groups based on their divergence at a lower level of similarity than the phenon line. We show that the identification of T. cruzi and T. rangeli in naturally mixed infections is readily achieved by either RAPD or isoenzyme analysis.
Cilia‐Mediated Oriented Chemokinesis in Tetrahymena thermophila
Authors: LEICK VAGN , KOPPELHUS UFFE , ROSENBERG JENS
Published: May 2007

The role of the cilia in the locomotion (“gliding”) of Tetrahymena thermophila in a semi‐solid medium has been studied when cells were migrating in gradients of attractant. Video recordings and computer‐aided motion analysis of migrating cells and their ciliary activity show that Tetrahymena thermophila migrate by swimming forward in semi‐solid methyl cellulose, using their cilia. Ciliary reversals occur at certain intervals and cause a termination (“stop”) of cellular migration. Cells with reversed cilia resume forward migration when normal ciliary beating resumes. In gradients of attractants, cells migrating towards the attractant suppress ciliary reversals, which leads to longer runs between stops than in control cells. Cells migrating away from the attractant have a higher frequency of ciliary reversals than the control cells resulting in shorter runs. Stimulated cells adapt to a particular ambient concentration of attractant several times during migration in the gradient. Adaptation is followed by de‐adaptation, which occurs during the “stop”. In the presence of cycloheximide, a strong inhibitor of chemoattraction, the attractant‐induced suppression of ciliary reversal is abolished (cells become desensitized to the attractant). It is concluded that Tetrahymena has a short‐term memory during adaptation. This is important for the efficiency of migration towards an attractant.
The Role of Trichocyst Discharge and Backward Swimming in Escaping Behavior of Paramecium from Dileptus margaritifer1
Authors: HARUMOTO TERUE
Published: May 2007

Trichocyst discharge is an effective defense of Paramecium against Dileptus margaritifer. The possible defensive function of backward swimming, which often follows trichocyst discharge upon Paramecium‐Dileptus encounters was studied. Mutants incapable of backward swimming (pawnA in P. tetraurelia, cnrA in P. caudatum) escaped from dilepti nearly as frequently as wild‐type cells. Double mutants (pawnA‐nd7, cnrA‐tnd2) were eaten nearly as frequently as mutants incapable of trichocyst discharge. Thus, in the defense of Paramecium against D. margaritifer, the role of backward swimming is minor, if any, compared to trichocyst discharge. Among escaped cells, about a half of wild‐type and essentially none of pawnA (cnrA) cells showed backward swimming. Paramecium behavior during the encounter can be mimicked by the local, not global, application of lysozyme which is a strong secretagogue of trichocyst.
Characterization of the Eyespot Regions of “Blind”Chlamydomonas Mutants after Restoration of Photophobic Responses
Authors: LAWSON MOIRA A. , SATIR PETER
Published: May 2007

Chlamydomonas reinhardtii exhibits photophobic and positive and negative phototactic responses that can be defined for cell populations using computerized cell tracking and motion analysis. Mutants CC‐2359 and FN68 are pigment deficient mutants that are blocked in carotenoid synthesis and lack these photo responses. In particular, neither mutant exhibits flash‐induced photophobic responses to visible light stimuli to which wild‐type gametic cells exhibit a strong response, with several behavioral stages. Upon addition of all‐trans retinal to these mutants, the photophobic responses are restored with minor quantitative differences from wild‐type populations. Using both light and electron microscopy, we have compared the ultrastructural characteristics of wild‐type C. reinhardtii to those of both mutants. As previously described, wild‐type cells contain an eyespot consisting of 2–4 layers of pigmented granules encased within thylakoid membranes, located between the distal extremities of the flagellar root. This structure is also visible as an orange‐red spot in light microscopy. The photoreceptor is thought to be concentrated in the plasma membrane above the eyespot. The mutant, CC‐2359, lacks this eyespot as seen by both light and electron microscopy, even when the photophobic response has been restored. FN68‐like mutants studied earlier by Morel‐Laurens and Feinlieb and others contain an eyespot which can be seen only by electron microscopy. In FN‐68, the eyespot generally has the same dimensions as in wt cells, differing mainly in pigment granule appearance. Consistent with these findings, several laboratories have shown that the full range of phototactic responses can be reconstituted in FN68 and CC‐2359, but that negative phototaxis requires a significantly stronger light stimulus in the latter strain. We confirm the suggestion that the eyespot is not necessary for the photophobic response, and is not critical for the appropriate assembly and function of the photophobic response receptor in the membrane. Furthermore, the locus of reconstitution of the functional receptor is not the eyespot. Because of the definitive demonstration of the absence of the eyespot in CC‐2359, however, the eyespot may play a role in negative phototaxis.
A New rDNA Repeat Unit in Human Giardia
Authors: UPCROFT JACQUELINE A. , HEALEY ANDREW , UPCROFT PETER
Published: May 2007

The rDNA repeat unit from a new human Giardia duodenalis strain shows significant differences from the previously reported G. duodenalis rDNA repeat. Twelve base‐pair changes occurred in 490 bp of the SSrRNA gene and new restriction enzyme sites occurred in the LSrRNA gene. The overall length of the rRNA genes is the same but the spacer is 76 bp longer than previously reported. A boundary within the spacers of the two different rDNA units divides a region of 50% homology near the LSrRNA gene from a region of 80% homology toward the SSrRNA gene. This boundary region includes two copies of a 78 bp repeat.
Evidence for the Ancestral Origin of Group I Introns in the SSUrDNA of Naegleria spp.
Authors: JONCKHEERE JOHAN F. DE
Published: May 2007

ABSTRACT The sequence variation within the group I intron in five Naegleria spp. was studied and compared with the sequence variation within the flanking small subunit ribosomal DNA. Considerable sequence divergence was observed in the introns as well as in the rDNA. In the intron deletions and insertions are only detected in the sequence contributing to the secondary structure, not in the open reading frame. Most of the sequence variation is detected in the unpaired loops. In the case of nucleotide substitution in helices, compensating base pair changes were observed. The sequence variation does not induce variation in the secondary structure model. The phylogenetic tree based on the intron sequences is similar to the tree based on the flanking rDNA sequences. This observation indicates that the intron might have been acquired at an early stage in evolution, and lost in the majority of Naegleria spp.
Revision of the Genus Protostelium (Eumycetozoa) I: The Protostelium mycophaga Group and the P. irregularis Group
Authors: SPIEGEL FREDERICK W. , GECKS SALLY C. , FELDMAN JOYCE
Published: May 2007

ABSTRACT The most commonly encountered protostelids are nominal members of the genus Protostelium Olive & Stoianovitch. These are Protostelium mycophaga Olive & Stoianovitch and P. irregularis Olive & Stoianovitch. Both species share the common features of long‐stalked fruiting bodies with single, uninucleate, deciduous spores and trophic states that consist solely of uninucleate amoebae, but they are quite different with respect to their detailed structure at both the light and electron microscopic levels. Based on this evidence, it seems unlikely that the species are congeneric, and it is proposed that P. irregularis be assigned to Soliformovum n. g. Examination of other species of Protostelium indicates that P. nocturnum Spiegel should be retained in the genus and that P. expulsum Olive & Stoianovitch should be reassigned to Soliformovum. The group most closely related to Protostelium, under this new concept, is the genus Planoprotostelium Olive & Stoianovitch, while Soliformovum is best treated as a genus of Eumycetozoa incertae sedis, though it does share some similarities with the group that includes the genera Ceratiomyxella Olive & Stoianovitch, Nematostelium Olive & Stoianovitch, and Schizoplasmodium Olive & Stoianovitch.
Isolation of the Flagellar Swelling and Identification of Retinal in the Phototactic Flagellate, Ochromonas danica (Chrysophyceae)
Authors: WALNE PATRICIA L. , PASSARELLI VINCENZO , LENZI PAOLA , BARSANTI LAURA , GUALTIERI PAOLO
Published: May 2007

Ochromonas danica, a freshwater, planktonic chrysophyte, is capable of sensing the light conditions of its environment. This biflagellate alga has a swelling near the base of the short flagellum and a chloroplastidic stigma in close association with it. A procedure is described for the isolation of this three dimensional flagellar swelling, the presumed photoreceptor. In contrast to an earlier method developed for the isolation of the paraflagellar swelling from Euglena gracilis, the protocol reported here for Ochromonas results in higher yields that should facilitate future biochemical investigations and could open avenues of investigation for the isolation and purification of the presumptive receptor protein. To verify the hypothesis that a rhodopsin‐like protein might be present in this alga, we applied a standard extraction procedure successfully used in the identification of retinal. We here report the purification and identification of all‐trans retinal in Ochromonas cells by column chromatography, HPLC and GC‐MS. Since retinal is the chromophore of rhodopsin‐like proteins, this finding may suggest that in these unicellular algae, too, a rhodopsin‐like protein could be the photoreceptor pigment.
Transcription Factor Genes from Rat Pneumocystis carinii
Authors: SUNKIN SUSAN M. , STRINGER JAMES R.
Published: May 2007

Genes encoding the TFIID TATA‐box binding protein (TBP) from two probable species of rat Pneumocystis carinii (prototype and variant) were sequenced. The two P. carinii TBP gene sequences were 91% identical to each other, and 65‐77% identical to TBP genes from other species. A cDNA from one of the two P. carinii TBP genes was sequenced, which showed that four small introns resided in identical positions within the TBP genes from the prototype and variant rat P. carinii. Conservation of the 180 amino acids that constitute the conserved core of TBP was 97% between the P. carinii TBP, which were 95% and 97% identical to conserved core sequences of TBP from Saccharomyces cerevisiae and Schizosaccharomyces pombe respectively.
Comparison of Coding and Spacer Region Sequences of Chromosomal rRNA‐Coding Genes of Two Sequevars of Pneumocystis carinii
Authors: ORTIZ‐RIVERA MARIA , LIU YONG , FELDER REGINA , LEIBOWITZ MICHAEL J.
Published: May 2007

Two distinct sequevars, denoted Pc1 and Pc2, of the opportunistic pathogen Pneumocystis carinii have been previously identified based on the sequence of their 26S rRNA genes, the location of group I self‐splicing introns and pulsed field electrophoretic patterns of chromosomal DNA. This study shows that the sequences of 16S and 5.8S rRNA genes also vary between these sequevars, and that greater variation was seen in the internal transcribed spacer regions. Polymerase chain reaction and restriction analysis can distinguish between these sequevars.
A Redescription of the Morphology of Onychodromus grandis Stein 1859 and the Systematic Implications of its Morphogenesis
Authors: SZABÓ ANDRÁS , WILBERT NORBERT
Published: May 2007

Morphology and morphogenesis of Onychodromus grandis (Hypotrichida, Ciliata) were studied in vivo and in protargolimpregnated preparations. The species was isolated from halophyte communities in astatic shallow waters of the Hortobágy National Park, Hungary. The species has two well developed dorsal horns. According to some characteristics (number and situation of ventral cirri) our findings differ from Stein's description in 1859. Individuals of the population studied had 9 ventral cirri in 3 rows. Nine rows of dorsal kineties were found on the back. It was found that the oral primordium develops in the vicinity of the transverse cirri. In O. grandis marginal primordia are formed in the marginal rows with the participation of parental cirri. This is also true for Laurentiella acuminata. Dorsal fragmentation of O. grandis displays similarity to that of L. acuminata, considering especially the development of the numerous small dorso‐marginal kineties. There are certain similarities among O. quadricornutus, O. indica, L. acuminata, Stylonychia vorax and O. grandis in the development of the ventral and dorsal pattern and in some morphogenetical events. The remarkable differences in the interphase cells can be related simply to the significantly different numbers of cortical units (number of cirri). and also to the various cell sizes.
Phylogenetic Relationships Among Anuran Trypanosomes as Revealed by Riboprinting
Authors: CLARK C. GRAHAM , MARTIN DONALD S. , DIAMOND LOUIS S.
Published: May 2007

Twenty trypanosome isolates from Anura (frogs and toads) assigned to several species were characterized by riboprinting–restriction enzyme digestion of polymerase chain reaction amplified small subunit ribosomal RNA genes. Restriction site polymorphisms allowed distinction of all the recognized species and no intraspecific variation in riboprint patterns was detected. Phylogenetic reconstruction using parsimony and distance estimates based on restriction fragment comigration showed Trypanosoma chattoni to be only distantly related to the other species, white T. ranarum and T. fallisi appear to be sister taxa despite showing non‐overlapping host specificities.
Phylogenetic Relationships of the Nassulida Within the Phylum Ciliophora Inferred from the Complete Small Subunit rRNA Gene Sequences of Furgasonia blochmanni, Obertrumia georgiana, and Pseudomicrothorax dubius
Authors: BERNHARD DETLEF , LEIPE DETLEF D. , SOGIN MITCHELL L. , SCHLEGEL K. MARTIN
Published: May 2007

ABSTRACT. Using comparisons of complete small subunit rRNA sequences from the ciliated protozoans Furgasonia blochmanni, Obertrumia georgiana, and Pseudomicrothorax dubius we inferred the phylogenetic position of the Nassulida (Class Nassophorea) within the Ciliophora. In distance matrix analyses the Nassulida share a common ancestry with the colpodean ciliate Colpoda inflata. Distance matrix and parsimony methods convincingly demonstrate that the Nassulida plus Colpodida are members of a complex ciliate assemblage that also includes the oligohymenophorans and phyllopharyngeans. These phylogenetic inferences are largely congruent with recent analyses of 23S‐like rRNA gene sequences and morphogenetic features. Groups traditionally thought to represent ancestral lineages now appear as highly derived ciliates. In contrast, heterotrichs which were considered to represent a highly evolved group, diverge at the base of the ciliates.
Giardia lamblia: Freeze‐fracture Ultrastructure of the Ventral Disc Plasma Membrane
Authors: CHAVEZ BIBIANA , MARTINEZ‐PALOMO ADOLFO
Published: May 2007

ABSTRACT. The close contact of Giardia lamblia trophozoites with mucosal surfaces produces surface indentations of epithelial cells, that progress to areas of microvilli depletion. This interaction is mediated by the lateral crest, a specialized contractile structure in the ventral disc of the parasite. We have analyzed the plasma membrane of the ventral disc using freeze fracture electron microscopy to study the distribution of large integral proteins, and freeze fracture cytochemistry with filipin to reveal sterol‐containing sites. A previously undetected structural specialization was found at the lateral crest as a horseshoe‐shaped membrane domain characterized by a near absence of intramembrane particles, in sharp contrast to the remainder of the ventral disc plasma membrane. In addition, the lateral crest showed a striking paucity of cholesterol‐containing complexes in replicas of trophozoites treated with filipin. These observations demonstrate that the plasma membrane of G. lamblia displays a microheterogeneity in the planar distribution of cholesterol and intramembrane particles. Both membrane components are noticeably less abundant at the outer rim of the ventral disc where contraction takes place.
A Cell Culture System for Study of the Development of Toxoplasma gondii Bradyzoites
Authors: WEISS LOUIS M. , LAPLACE DENISE , TAKVORIAN PETER M. , TANOWITZ HERBERT B. , CALI ANN , WITTNER MURRAY
Published: May 2007

ABSTRACT. Toxoplasma gondii is a ubiquitous apicomplexan parasite and a major opportunistic pathogen under AIDS‐induced conditions, where it causes encephalitis when the bradyzoite (cyst) stage is reactivated. A bradyzoite‐specific Mab, 74.1.8, reacting with a 28 kDa antigen, was used to study bradyzoite development in vitro by immuno‐electron microscopy and immunofluorescence in human fibroblasts infected with ME49 strain T. gondii. Bradyzoites were detected in tissue culture within 3 days of infection. Free floating cyst‐like structures were also identified. Western blotting demonstrated the expression of bradyzoite antigens in these free‐floating cysts as well as in the monolayer. Bradyzoite development was increased by using media adjusted to pH 6.8 or 8.2. The addition of γ‐interferon at day 3 of culture while decreasing the total number of cysts formed prevented tachyzoite overgrowth and enabled study of in vitro bradyzoites for up to 25 days. The addition of IL‐6 increased the number of cysts released into the medium and increased the number of cysts formed at pH 7.2. Confirmation of bradyzoite development in vitro was provided by electron microscopy. It is possible that the induction of an acute phase response in the host cell may be important for bradyzoite differentiation. This system should allow further studies on the effect of various agents on the development of bradyzoites.
Vittaforma corneae N. Comb. for the Human Microsporidium Nosema corneum Shadduck, Meccoli, Davis & Font, 1990, Based on its Ultrastructure in the Liver of Experimentally Infected Athymic Mice
Authors: SILVEIRA HENRIQUE , CANNING ELIZABETH U.
Published: May 2007

ABSTRACT. A new genus, Vittaforma n. g. is proposed for the human microsporidium Nosema corneum Shadduck, Meccoli, Davis & Font, 1990, based on the ultrastructure of developmental stages in the liver of experimentally infected athymic mice. The diplokaryotic arrangement of the nuclei is the only character that conforms with the description of the genus Nosema. Sporogony is polysporoblastic, sporonts are ribbon‐shaped, constricting to give rise to linear arrays of sporoblasts and each parasite is enveloped by a complete cistema of host endoplasmic reticulum. Comparison of N. corneum, with established genera revealed that there were none with the same combination of characters. Consequently it is proposed that Nosema corneum be placed in a new genus as Vittaforma corneae n. comb.
Conjugation Rescue of Exocytosis Mutants in Tetrahymena thermophila Indicates the Presence of Functional Intermediates in the Regulated Secretory Pathway
Authors: SAUER MONICA K. , KELLY REGIS B.
Published: May 2007

ABSTRACT. Tetrahymena thermophila possesses a regulated secretory pathway in which mucin proteins are stored in dense‐core granules, called mucocysts. Exocytosis‐defective mutants exist that fail to secrete mucin in response to secretagogues. Four of the mutants (SB281, SB283, SB285 and SB715) appear to be blocked at different steps of the regulated secretory pathway. SB281 and SB285 accumulate mucin proteins in heterogeneous cytoplasmic organelles which have not yet been identified; SB283 makes mucocyst‐like structures but they contain no immunologically identifiable 80‐kDa or 50‐kDa mucin proteins; and SB715 has more than normal amounts of immature and undocked mucocysts. The organelles that accumulate in exocytosis‐defective mutants could be either normal intermediates in the biosynthetic pathway or aberrant structures that form as a result of the mutations. We have used conjugation rescue to analyze steps in the biogenesis of exocytosis‐competent mucocysts and to identify functional intermediates. The cytoplasmic organelles that accumulate in SB281 appear to be unidentified biosynthetic intermediates, and the defect is in a cytosolic protein essential for mucocyst maturation. The organelles which accumulate in the other mutants are likely biosynthetic, but their mutations are in proteins which are labile or not free to diffuse into the mutant conjugant.
Comparison of Methods of Cell Cycle Analysis in Paramecium tetraurelia
Authors: ADL SINA M. , BERGER JAMES D.
Published: May 2007

ABSTRACT. Four methods are commonly used to study cell cycle processes in Paramecium tetraurelia. These include stage frequency analysis in asynchronous cultures, hand selection of synchronous dividing cells, selection of newly divided cells by elutriation centrifugation, and the sister cell method. We have compared the timing and resolution of stages of oral morphogenesis and micronuclear mitosis with each method. The temporal resolution obtainable with the sister cell method was inadequate to position the timing of morphogenesis stages within the cell cycle. Both the asynchronous method and the hand‐selected synchronous samples methods are prone to bias. Elutriation centrifuge synchronization provides large samples with resolution comparable to that of hand selected samples. The elutriation method is the least prone to bias when <5% of the parent culture of Paramecium is selected.
Ultrastructure and Rapid Axopodial Contraction of a Heliozoa, Raphidiophrys contractilis Sp. Nov.
Authors: KINOSHITA EIJI , SUZAKI TOSHINOBU , SHIGENAKA YOSHINOBU , SUGIYAMA MASANORI
Published: May 2007

ABSTRACT. In the present study, we isolated a species of heliozoans from a brackish pond in Shukkeien Garden, Naka‐ku, Hiroshima City, Japan. Electron‐microscopic observations showed that the axonemal microtubules in this heliozoan constituted a complex pattern of hexagons and triangles. By applying SDS‐polyacrylamide gel electrophoresis and subsequent immunoblotting, molecular weights of α and β‐tubulins were determined to be 48 and 45 kDa, respectively. X‐ray microanalysis demonstrated that the numerous scales coating the cell body surface were silicic structures. Size and shape of the cell body and the scales were examined and compared with other known species of heliozoans, which led us to conclude that this is a new species belonging to the genus Raphidiophrys. This heliozoan was also found to carry out rapid axopodial contraction during food uptake at a velocity of about 1 mm/s. With reference to this characteristic contractile behavior, this new species was named Raphidiophrys contractilis.
Cloning and Characterization of an ATPase Gene from Pneumocystis carinii which Closely Resembles Fungal H+ ATPases
Authors: MEADE JOHN C. , STRINGER JAMES R.
Published: May 2007

ABSTRACT. A gene encoding a P‐type cation translocating ATPase was cloned from a genomic library of rat‐derived Pneumocystis carinii. The nucleotide sequence of the gene contains a 2781 base‐pair open reading frame that is predicted to encode a 101, 401 dalton protein composed of 927 amino acids. The P. carinii ATPase protein (pcal) is 69–75% identical when compared with eight proton pumps from six fungal species. The Pneumocystis ATPase is less than 34% identical to ATPase proteins from protozoans, vertebrates or the Ca++ ATPases of yeast. The P. carinii ATPase contains 115 of 121 residues previously identified as characteristic of H+ ATPases. Alignment of the Pneumocystis and fungal proton pumps reveals five homologous domains specific for fungal H+ ATPases.
Effect of Fetal Bovine Serum Glycoproteins on the In Vitro Proliferation of the Oyster Parasite Perkinsus marinus: Development of a Fully Defined Medium
Authors: GAUTHIER JULIE D. , FEIG BEVERLY , VASTA GERARDO R.
Published: May 2007

ABSTRACT. The oyster parasite Perkinsus marinus replicates in our medium consisting of Dulbecco modified Eagle's medium: Ham's F12 nutrient mixture (1:1) supplemented with 1–5% fetal bovine serum, with a doubling time of 24 hours during the exponential phase of the culture. Fetal bovine serum concentrations above 5% dramatically reduced parasite proliferation in a dose‐dependent manner. We tested the individual effects of the three major protein components of fetal bovine serum (fetuin, transferrin and albumin) on the replication of the parasite in a serum‐free medium. At the concentrations tested, fetuin enhanced parasite growth, whereas albumin had a modest positive effect and transferrin was inhibitory. Proteolytic digestion of fetuin, strongly diminished its growth‐enhancing properties, indicating that the overall glycoprotein architecture may be required for activity. On the contrary, desialylation of fetuin slightly enhanced its growth‐promoting activity. The addition of fetuin at 1.7 mg/ml to the serum‐free DME: Ham's F12 medium yielded growth rates that are comparable to those obtained with our standard culture methodology. This has resulted in a fully defined culture medium that will allow for a rigorous characterization of excretory/secretory products involved in modulating or blocking the host's humoral and cellular defense mechanisms.
In Vitro Culture of Phytomonas Sp. Isolated from Euphorbia characias. Metabolic Studies by 1H NMR
Authors: SÁNCHEZ‐MORENO MANUEL , FERNANDEZ‐BECERRA CARMEN , ENTRALA EMILIO , OPPERDOES FRED R. , DOLLET MICHEL , OSUNA ANTONIO
Published: May 2007

ABSTRACT. We describe the in vitro culture of Phytomonas species isolated from Euphorbia characias. The best choice between tested media was SDM‐79, in which promastigotes, after 6 days of culture, reached cell densities as high as 4 × 107 cells/ml. Cells growing in LIT or MTL medium showed longer division times and lower cell densities. We succeeded in obtaining Phytomonas sp. amastigote and spheromastigote forms in modified GRACE's medium, yielding transformation rates of up to 70%. Electron microscopy studies were performed in order to characterize the ultrastructural features of these forms obtained in vitro. On the other hand, metabolic studies based on qualitative (nuclear magnetic resonance spectroscopy) and quantitative metabolic methods (enzymatic assays) showed that promastigote forms secreted mainly ethanol, acetate, glycine, glycerol, piruvate and succinate in SDM‐79 medium, whereas the major metabolites found after transformation in modified Grace's medium were ethanol, acetate, glycine, piruvate and smaller amounts of glycerol.
Towards a Non‐Terminal Viability Assay for Foraminiferan Protists
Authors: BERNHARD JOAN M. , NEWKIRK SARAH G. , BOWSER SAMUEL S.
Published: May 2007

ABSTRACT. Epifluorescence microscopy and spectrofluorimetry were investigated as possible non‐terminal methods to distinguish live from dead foraminifera. Seven fluorogenic probes (diacetates of fluorescein [FDA], carboxyfluorescein, dichlorofluorescein, and carboxyeosin; AM‐esters of biscarboxyethylcarboxyfluorescein [BCECF‐AM], calcein, and calcein blue) were tested on Allogromia laticollaris. The probes that consistently produced the brightest fluorescence signals (BCECF‐AM and FDA) were judged non‐toxic to Allogromia, on the basis of short‐term pseudopodial deployment and long‐term reproduction assays. Once protocols were established, these two probes were tested on 13 additional benthic foraminiferal species. We found that BCECF‐AM is the most suitable probe for direct epifluorescence microscopy of metabolically active foraminifera, especially tectinous and transparent calcareous species. Using spectrofluorimetry, FDA showed promise for opaque species because fluorescence is detected in the incubation media after its release from the cell. However, both approaches could only be used with confidence in light of appropriate controls established for each species examined.
Mono‐ADP‐Ribosylation of Arginine Residue of Euglena gracilis Z in Synchronous Culture
Authors: TAKENAK SHIGEO , INAGAKI JUNKO , TSUYAMA SHINGO , MIYATAKE KAZUTAKA , NAKANO YOSHIHISA
Published: May 2007

ABSTRACT. In Euglena gracilis Z, a considerably high activity of mono‐ADP‐ribosyltransferase occurred and change of it was accompanied by a cell cycle induced by a light‐dark cycle. The enzyme activity was strongly inhibited by L‐arginine and supported in the presence of poly‐L‐arginine as a substrate, indicating that ADP‐ribosylated amino acid is an arginine residue. Arginine: mono‐ADP‐ribosyltransferase activity was found in the chloroplasts, mitochondria, microsomes and cytosol as judged from marker enzyme activities and the activity in each organelle fluctuated with the cell cycle.
Evidence of a Cadmium‐Thionein and the Glycine Cleavage System in Oxytricha granulifera
Authors: IRATO PAOLA , PICCINNI ESTER , JAMES PETER , AMMERMANN DIETER
Published: May 2007

ABSTRACT. This work presents the further purification of a Cd‐linking protein in Oxytricha granulifera by reverse‐phase chromatography. This protein contains 25% cysteine and no aromatic amino acid. it may be considered as a chelatin with some similarity to metallothioneins. During the purification of another Cd‐linking compound, we were able to demonstrate that the H protein precursor of glycine cleavage is present in Oxytricha. This is the first finding of the presence of this system in Protozoa.
Culture and Phylogenetic Characterization of Trichomitus trypanoides Duboscq & Grassè 1924, n. comb.: a Trichomonad Flagellate Isolated from the Hindgut of the Termite Reticulitermes santonensis Feytaud
Authors: BERCHTOLD MANFRED , BREUNIG ALFRED , KÖNIG HELMUT
Published: May 2007

ABSTRACT. A trichomonad flagellate strain R1 was isolated from the hindgut contents of the termite Reticulitermes santonensis Feytaud. The flagellate was cultivated at 28° C in anaerobic medium containing yeast extract, minerals and vitamins. The isolate fed on living bacteria. It showed the typical morphological and ultrastructural features of the trichomonads. closely resembling Trichomitus trypanoides. In order to determine its phylogenetic position the small subunit ribosomal DNA (SSU rDNA) of the flagellate was amplified in vitro using the polymerase chain reaction (PCR), cloned in a plasmid vector and sequenced. Comparison of the obtained sequence with so far available SSU rRNA/rDNA sequences showed strongest similarity (89%) to the sequence of Tritrichomonas foetus. The phylogenetic analysis with parsimony and distance matrix methods placed Trichomitus trypanoides strain R1 near by the root of the phylogenetically so far analyzed eukaryotic organisms. This confirms that termites harbour hindgut symbionts, which originate from very early evolved eukaryotes.
Two New Menosporine Gregarines, Hoplorhynchus acanthatholius N. Sp. and Steganorhynchus dunwoodyi N. G., N. Sp. (Apicomplexa: Eugregarinorida: Actinocephalidae) from Coenagrionid Damselflies (Odonata: Zygoptera)
Authors: PERCIVAL TAMARA J. , CLOPTON RICHARD E. , JANOVY JOHN
Published: May 2007

ABSTRACT. Hoplorhynchus acanthatholius, n. sp. is described from Enallagma civile, the Civil Bluet damselfly. Trophozoites are solitary, lie in the mesenteron between the peritrophic membrane and the epithelium, and attain a maximum length of 850 μm. Epimerite ovoid to broadly ovoid; anterior margin bearing eight equidistant retroarcuate hooks; attached to protomerite by means of a vermicular stalk. Protomerite ovoid; deutomerite narrowly obvoid. Gametocysts spherical; diam 300 μm, sporulating by simple dehiscence in 48–72 h. Oocysts are characteristic of Menosporinae: smooth, biconical, crcscentic, uniform in size and shape. Steganorhynchus dunwoodyi, n. g., n. sp. is described from the damselfly Ischnura verticalis. The genus is characterized by an epimerite comprising an ovoid papilla enclosed in a retractable, globular sheath, borne on a long vermicular stalk. Trophozoites are solitary, lie in the mesenteron between the peritrophic membrane and epithelium, and attain a maximum length of 605 μm. Protomerite very broadly ovoid; deutomerite obvoid. Gametocysts spherical; diam 258 μm, sporulating by simple dehiscence in 48–72 h. Oocysts are characteristic of Menosporinae: smooth, biconical, crescentic, uniform in size and shape. The population dynamics of H. acanthatholius and S. dunwoodyi among damselfly populations in five Nebraska localities are presented.
Coordinated Flagellar and Ciliary Beating in the Protozoon Tritrichomonas foetus1
Authors: MONTEIRO‐LEAL LUIZ H. , FARINA MARCOS , BENCHIMOL MARLENE , KACHAR BECHARA , DE SOUZA WANDERLEY
Published: May 2007

ABSTRACT. Tritrichomonas foetus is a flagellated protozoon found in urogenital tract of cattle. Its free movement in liquid medium is powered by the coordinated movement of three flagella projecting towards the anterior region of the cell, and one recurrent flagellum that forms a junction with the cell body and ends as a free projection in the posterior region of the cell. We have used video microscopy and digital image processing to analyze the relationships between the movements of these flagella. The anterior flagella beat in a ciliary type pattern displaying effective and recovery strokes, while the recurrent flagellum beats in a typical flagellar wave form. One of the three anterior flagella has a distinctive pattern of beating. It beats straight in its forward direction as opposed to the ample beats performed by the others. Frequency measurements obtained from cells swimming in a viscous medium shows that the beating frequency of the recurrent flagelium is approximate twice the frequency for the three anterior flagella. We also observed that the costa and the axostyle do not show any active motion. On the contrary, they form a cytoskeletal base for the anchoring and orientation of the flagella.
A Simple, Efficient Technique for Freezing Tetrahymena thermophila
Authors: CASSIDY‐HANLEY DONNA , SMITH HELEN R. , BRUNS PETER J.
Published: May 2007

ABSTRACT. We have developed a simple, efficient procedure for the long term freezing of Tetrahymena thermophila in liquid nitrogen. This technique yields excellent recovery of viable cells with all strains tested and does not require the use of a controlled rate low temperature freezer. To optimize the freezing technique, we have examined the effects of varying a number of parameters, including the physiological state of the cells prior to freezing, the time of exposure to cryoprotectant, and the rate of freezing and thawing. the frequency of viable cell recovery following freezing using this technique has been tested for a variety of different cell lines.
Effects of pH on Excystation of Gregarina cuneata and Gregarina polymorpha (Eugregarinida: Gregarinidae)1
Authors: CLOPTON RICHARD E. , GOLD ROGER E.
Published: May 2007

ABSTRACT. The mechanisms that signal gregarine excystation are unknown. Previous authors have demonstrated that gregarine sporozoites excyst from their surrounding oocyst in response to stimuli contained in host digestive fluids, but the role of host intestinal pH in this signaling system has not been investigated. In this study, an in vitro assay is used to quantify the effects of 3 pH levels (6.1, 7.0, 8.0) on the excystation of two gregarine species, Gregarina cuneata and Gregarina polymorpha. Both gregarine species excyst at all three pH levels, but there are significant within and among species differences in excystation rate and cumulative excystation over time. Gregarina cuneata excysts more rapidly at pH 6.0 and G. polymorpha excysts more rapidly at pH 8.0. Cumulative excystation is maximized at pH 6.0 for G. cuneata and at pH 7.0 for G. polymorpha. Hydrogen ion mediated excystation may lead to the formation of foci for subsequent establishment or migration and may play a role in parasite site specificity.
Analysis of the Soluble and Membrane‐bound Immobilization Antigens of Ichthyophthirius multifiliis
Authors: XU CHUNHUI , CLARK THEODORE G. , LEFF ADAM A. , DICKERSON HARRY W.
Published: May 2007

ABSTRACT. The immobilization antigens (i‐antigens) are a class of highly abundant surface membrane proteins found on a number of holotrich ciliates. In Ichthyophthirius multifiliis (an obligate parasite of fish) these antigens appear to be targets of the host immune response. While the i‐antigens of Ichthyophthirius are predominantly membrane‐associated proteins, we now find that they are released into the water surrounding the parasite in a highly enriched form. The membrane‐associated and water soluble proteins appear indistinguishable by antigenic means, as well as by several biochemical criteria including peptide mapping, mobility in reducing and non‐reducing SDS‐polyacrylamide gels, and relative glycosylation. Antibodies raised against the membrane‐associated antigens react with the water soluble proteins on Western blots. Not surprisingly, immunocytochemical localization studies show binding of these antibodies to surface membranes of the cell. In addition, however, antibody binding is also detectible on the membranes of a secretory organelle (that is, mucocysts) present in the cortical cytoplasm. The significance of these findings with regard to the potential role of the i‐antigens in infection and immunity is discussed.