bluuebunny/crossref_metadata_2025 huggingface dataset:
Evidence for a Viral Macronuclear Endosymbiont in Paramecium caudatum
Authors: TSUKII YUUJI , HARUMOTO TERUE , YAZAKI KAZUMORI
Published: May 2007

ABSTRACT. Some strains of P. caudatum contain macronuclear inclusion bodies that are morphologically distinct from bacteria. They vary in number as well as in size in each macronucleus. The inclusion bodies are basically divided into peripheral and inner areas. The peripheral area consists of fibrillar proteins of 22–24 nm in thickness, which are specifically stained with fast green in 45% acetic acid. On the other hand, chromatin‐like granules are within the inner area of large inclusion bodies. The granules within the inner area changed their distribution depending upon the physiological state of their host cells. Transplantation experiments and crossbreeding analyses revealed that genetic factors responsible for the multiplication of the inclusion bodies can ‘infect’ other macronuclei (or cells) via the cytoplasm. These results suggest that the inclusion bodies are a non‐bacterial macronuclear endosymbiont, possibly produced by a virus or a virus‐like element.
Metabolites of the Free‐Living Amoeba Phreatamoeba balamuthi Analyzed by 13C‐ and 31P‐NMR Spectroscopy: Occurrence of Phosphoinositol Diphosphates
Authors: MARTIN JEAN‐BAPTISTE , BAKKER‐GRUNWALD TILLY , KLEIN GERARD
Published: May 2007

ABSTRACT. Phreatamoeba balamuthi is a free‐living heterotrophic amoeba that lacks mitochondria. Metabolites of axenically‐grown cells were characterized by natural‐abundance 13C‐NMR and 31P‐NMR spectroscopy on acellular perchloric acid extracts. The amoebae were found to contain glycogen and trehalose as storage carbohydrates, together with putrescine and several amino acids, most prominently proline; we propose that proline and trehalose may serve in osmoregulation. Glycerophosphocholine and glycerophosphoethanolamine were present with their phosphomonoester derivatives, phosphocholine and phosphoethanolamine. Along with inorganic phosphate, inorganic pyrophosphate, nucleoside diphosphates, nucleoside triphosphates and NAD, P. balamuthi amoebae also contained unusual phosphoinositol diphosphates in large quantities (0.5 μmol/g wet cells).
Characterization of the Target Antigens of Phytomonas‐specific Monoclonal Antibodies
Authors: TEIXEIRA MARTA M. G. , CAMPANER MARTA , CAMARGO ERNEY P.
Published: May 2007

ABSTRACT. Seven Phytomonas‐specific monoclonal antibodies produced against Phytomonas serpens and Phytomonas françai were further characterised in order to identify and localise their target antigens. Four monoclonal antibodies recognized carbohydrate surface epitopes, in three of the cases associated with surface glycoproteins with apparent molecular weight of 80 kDa. One monoclonal antibody apparently bound to a surface/internal protein epitope, whereas the two others recognized intra‐cellular proteins. The cell surface epitopes recognized by monoclonal antibodies were detected specifically in the genus Phytomonas. These epitopes, which are detected in culture, plant and insect forms, may be useful as targets for Phytomonas identification.
The NADP‐linked Aldehyde Reductase of Leishmania donovani
Authors: KEEGAN FRANK P. , BLUM J. JOSEPH
Published: May 2007

ABSTRACT. An enzyme that oxidizes ethanol to acetaldehyde in the presence of NADP (but not NAD) and reduces acetaldehyde to ethanol in the presence of NADPH (but not NADH) is present in Leishmania donovani promastigotes. The activity is present only in the supernatant fraction obtained from sonication of the cells and high speed centrifugation. The Km and Vm values were evaluated for propanol and propionaldehyde as well as for ethanol and acetaldehyde in cells obtained from late log and 3‐day stationary phase cultures. There was no significant change in Km or Vm values for any of these four substrates with culture age. Since the Km values for ethanol and propanol are much higher than for the corresponding aldehydes and higher than any physiological range of alcohol concentration likely to be encountered, this enzyme is considered to function as an aldehyde reductase.
Modeling the Individual Growth of Tetrahymena Sp. and its Population Consequences
Authors: RATSAK CHRISTA H. , KOOI BOB W. , KOOIJMAN BAS
Published: May 2007

ABSTRACT. Three types of mathematical growth models are presented to describe the individual growth of the ciliate Tetrahymena sp. feeding on the bacterium Pseudomonas fluorescens. Both organisms were isolated from a domestic waste‐water treatment plant. Growth of individual ciliates and the consequences for the whole population are considered. Experimental data, obtained by following the individual ciliate during its lifespan from cell division to cell division, are used for parameter estimations. Differences between growth models for individuals turn out to have little effect on the specific population growth rate and the mean cell volume. In case of exponential growth of individuals the unstructured and structured population models are equivalent, even in time‐variant environments. This knowledge can be applied in the stability analysis of food chains or forced systems. The results obtained facilitates quantification of protozoa biomass as a function of bacterial biomass in chemostats. More specifically, it highlights the dynamic behaviour of bacteria and protozoa in waste‐water treatment plants.
Inhibition of Protein Synthesis and Amino Acid Transport by Crystal Violet in Trypanosoma cruzi
Authors: HOFFMANN MARIA E. , JANG JUNHYUK , MORENO SILVIA N. J. , DOCAMPO ROBERTO
Published: May 2007

ABSTRACT. [35S]methionine incorporation into proteins of either T. cruzi epimastigotes or trypomastigotes was drastically inhibited by low concentrations of crystal violet in a dose‐dependent manner. This inhibition was not due to ATP depletion since cellular ATP levels did not change significantly after incubation of epimastigotes with 50 μM crystal violet for similar periods of time, and was unaffected by changes in the extracellular free calcium concentration. Although crystal violet was able to inhibit protein synthesis in a cell‐free system from T. cruzi epimastigotes, half maximal inhibition was at 1 mM, a concentration three orders of magnitude higher than those that inhibited protein synthesis in intact cells. On the other hand, crystal violet was able to inhibit total [35S]methionine uptake at similar concentrations to those that inhibited protein synthesis while addition of increasing concentrations of cold methionine to the incubation medium protected the cells against crystal violet inhibition. Crystal violet also inhibited total [3H]proline uptake thus indicating that it has a general inhibitory effect upon the transport of amino acids, and not specifically upon methionine. These results indicate that inhibition of protein synthesis by crystal violet is probably due to inhibition of amino acid uptake.
Comparison of the 24 kDa Flagellar Calcium‐Binding Protein cDNA of Two Strains of Trypanosoma cruzi
Authors: GODSEL LISA M. , OLSON CHERYL L. , LACAVA ZULMIRA G. M. , ENGMAN DAVID M.
Published: May 2007

ABSTRACT. DNA sequences encoding the 24 kDa flagellar calcium binding protein (FCaBP) of two strains of Trypanosoma cruzi were found to differ at fourteen positions, six of which result in amino acid differences. Four of the amino acid differences are located within the calcium‐binding domains of FCaBP; however, none is predicted to affect the calcium‐binding ability of the protein. Chromosomes harboring the FCaBP gene clusters differ in size among T. cruzi strains.
A Three Nucleotide Signature Sequence in Small Subunit rRNA Divides Human Giardia in Two Different Genotypes
Authors: VAN KEULEN HARRY , HOMAN WIEGER L. , ERLANDSEN STANLEY L. , JARROLL EDWARD L.
Published: May 2007

ABSTRACT. The nucleotide sequence of the 16S rRNA gene, part of the 23S rRNA gene and the spacer DNA region was determined for Giardia duodenalis, obtained from humans in The Netherlands (AMC‐4) and Washington State (CM). These rDNA sequences differ from other G. duodenalis isolates (Portland‐1 and BRIS/83/HEPU/106) both of which have virtually identical rDNA sequences. The most characteristic feature was found close to the 5’end of the 16S rRNA. The Portland‐1 ‐ Bris/83/HEPU/106 type has GCG in position 22–24, while AMC‐4 and CM have AUC in this position. These two sequences, present in an otherwise conserved region of the 16S rRNA, are “signature” sequences, which divide Giardia isolates into two different groups.
Proteolysis of a 34 kDa Phosphoprotein Coincident with a Decrease in Protein Kinase Activity During the Erythrocytic Schizont Stage of the Malaria Parasite
Authors: WISER MARK F.
Published: May 2007

ABSTRACT. Protein phosphorylation events may play important roles in the replication and differentiation of the malarial parasite. Investigations into the lability of a Plasmodium protein kinase revealed that a 34 kDa parasite phosphoprotein is rapidly converted into a 19 kDa fragment. Coincident with this conversion is a nearly total loss of a protein kinase activity, as determined from the phosphorylation of endogenous protein substrates. Both the conversion of the 34 kDa protein to the 19 kDa protein and the loss of protein kinase activity are inhibited by thio‐protease inhibitors. The presence of low levels of the intact 34 kDa protein restores the protein kinase activity to almost maximum levels. However, it was not possible to demonstrate protein kinase activity associated with the 34 kDa protein, thus suggesting that the 34 kDa protein is probably an activator or regulator of the protein kinase activity and not a protein kinase. The conversion to the 19 kDa fragment also occurs in vivo and only during the schizont stage prior to the appearance of ring forms. During this same period the protein kinase activity decreases suggesting that the proteolytic processing of the 34 kDa protein may be a physiological regulator of the protein kinase.
Analysis of Genetic Diversity at the arom Locus in Isolates of Pneumocystis carinii
Authors: BANERJI SUNEALE , LUGLI ELENA B. , MILLER ROBERT F. , WAKEFIELD ANN E.
Published: May 2007

ABSTRACT. The DNA sequences of a portion of the 5‐enolpyruvyl shikimate phosphate synthase domain of the arom gene, encoding the pentafunctional AROM protein, were determined from isolates of Pneumocystis carinii from five mammalian host species (rat, human, ferret, rabbit and mouse). High levels of genetic divergence were found among P. carinii derived from different hosts species, 7–22% at the DNA sequence level, and 7–26% at the derived amino acid sequence level. Two separate and distinct sequences were isolated from infected ferret lungs. Low levels of divergence were seen in human‐derived organisms.
Catabolism of Tryptophan by Trypanosoma evansi
Authors: EL SAWALHY AHMED , SEED JOHN R. , EL ATTAR HASSAM , HALL JAMES EDWIN
Published: May 2007

ABSTRACT. Trypanosoma brucei gambiense, which causes human African trypanosomiasis, catabolizes the aromatic amino acid tryptophan via an initial aminotransferase catalyzed reaction to form several indole end products, which have been suggested to contribute to the pathogenesis of trypanosomiasis. To determine if this same pathway exists in T. evansi, the closely related trypanosome pathogen of domestic animals, tryptophan catabolism was examined in vitro and in vivo. As is the case with human African trypanosomes, T. evansi catabolized tryptophan to form indole‐3‐pyruvic acid and smaller amounts of indole‐3‐acetic acid and indole‐3‐lactic acid. Large concentrations of indole‐3‐pyruvic acid are excreted in urine of trypanosome‐infected mice. However, indole‐3‐ethanol could not be detected in incubates of T. evansi or T. b. gambiense, even though the latter species had previously been reported to form this neutral metabolite. A new, previously unreported tryptophan metabolite was isolated and partially characterized from incubates of T. evansi and T. b. gambiense. Although the functional significance of tryptophan catabolism to trypanosomatids remains obscure, the pathway is quantitatively significant in all species examined thus far.
Comparison of Suitability of Great Bay, New Jersey, and Parsonage Creek, New York, for Alexandrium tamarense
Authors: MAHONEY JOHN B. , McGHEE JULIE A. , McNULTY J. KNEELAND
Published: May 2007

ABSTRACT. Alexandrium tamarense (Lebour) Balech (=Gonyaulax tamarensis Lebour) has been widely distributed and occasionally abundant in coastal waters of Long Island, New York in recent years. However, the distribution on the New Jersey coast has been sparse and this fact cannot be explained by this dinoflagellate's ability to migrate, or by its nutritional and physiological characteristics, or by the region's general suitability for phytoplankton. Therefore, the possibility that New Jersey coastal waters might be chemically exclusionary for A. tamarense seemed worth exploring. In a limited approach, we tested for water quality detrimental to the species in one New Jersey site (Great Bay) with a series of annual assays. Parsonage Creek, Long Island, New York, was assayed for comparison; this creek is assumed to have had at least marginal suitablity for A. tamarense, based on its reported long‐term presence. Results provide tentative support for our working hypothesis, i.e. Great Bay chemical water quality is generally unfavorable for A. tamarense. Inhibition of A. tamarense growth, or culture decline, occurred in both assay series, but was substantially greater in Great Bay water. Inimical water quality was the most important factor distinguishing the two sites. Chelation with EDTA had greatest overall benefit in Great Bay assays, suggesting that lower availability of a natural chelator in the bay could be a secondary factor. Assay metal response is problematic, but we believe it permits speculation that essential metals could be partially limiting to A. tamarense in Great Bay, but would not be a critical regulator.
Analysis of a Repeat‐Containing Family of Giardia lamblia Variant‐Specific Surface Protein Genes: Diversity Through Gene Duplication and Divergence
Authors: YANG YONG‐MIN , ADAM RODNEY D.
Published: May 2007

ABSTRACT. Giardia lamblia trophozoites express on their surfaces one of a set of cysteine‐rich antigenically variant proteins, called variant‐specific surface proteins, which comprise the majority of proteins detected by surface labeling. While these VSP proteins may be immunodominant proteins important in the host immune response to G. lamblia, the ability to switch expression from one VSP to another may provide a means for the trophozoites to avoid the host immune response. The first VSP characterized, VSPA6 (from the A6 clone of the WB isolate, originally termed CRP170), contains 18–23 copies of a 65 amino acid repeat. We have now used the repeat as a probe to isolate from a WBA6 genomic library two genes related to vspA6 (called vspA6‐S1, vspA6‐S2). Sequence analysis of the vspA6‐S1 gene revealed nearly two complete copies of the 195 bp repeat and substantial nucleotide and translated amino acid similarity in the coding regions 5’and 3’to the repeats. The vspA6‐S2 gene, while still related, showed greater divergence from vspA6 than vspA6‐S1 in the nonrepeat coding region and contained nearly four copies of a 201 bp repeat that was 75% identical to the 195 bp vspA6 repeat. These results suggest that gene duplication followed by divergence has played a key role in the generation of the vsp gene repertoire.
Transmissibility of Bacterial Endosymbionts Between Isolates of Acanthamoeba Spp.
Authors: GAUTOM ROMESH K. , FRITSCHE THOMAS R.
Published: May 2007

ABSTRACT. Experimental transmission of two bacterial endosymbionts to symbiont‐free isolates of Acanthamoeba spp. was studied to determine specificity of the host‐symbiont relationship. Both symbionts originated from amoebic isolates displaying an identical mitochondrial DNA EcoRI fingerprint (group AcUW II). Symbioses were readily established in one amoebic isolate which displayed a homologous mtDNA fingerprint (group AcUW II). Exposure of a heterologous amoebic isolate (group AcUW IV) to the two symbionts resulted in either cell death or encystation without the establishment of symbioses. While symbioses were established with an amoebic isolate from a second heterologous group (AcUW I), a unique membranous sheath appeared and persisted around one of the symbionts which did not exist in the original host. An isolate representing a third heterologous amoebic group (AcUW VI) was variable in its susceptibility with one symbiont unable to infect the host and the other becoming established only after an initial reaction in which trophozoites rounded‐up and floated off the substrate. These studies suggest that a specific recognition system exists between particular isolates of Acanthamoeba and their symbionts, and that the appearance of a killer phenotype is related to contact between mismatched, though recognized, pairs.
Molecular Comparison of the Mitochondrial and Cytoplasmic hsp 70 of Trypanosoma cruzi, Trypanosoma brucei and Leishmania major
Authors: KLEIN KARIN G. , OLSON CHERYL L. , DONELSON JOHN E. , ENGMAN DAVID M.
Published: May 2007

ABSTRACT. We compared the expression and localization of the mitochondrial and cytoplasmic hsp70 of the protozoans Trypanosoma cruzi, Trypanosoma brucei and Leishmania major. The mitochondrial protein is encoded by multiple mRNA in all species, while the cytoplasmic protein is encoded by a single mRNA. In all three species, the mitochondrial hsp70 is concentrated in the kinetoplast, a submitochondrial structure that houses the unusual DNA (kDNA) that characterizes this group of organisms, while the cytoplasmic protein is distributed throughout the cell. These results suggest that, in all kinetoplastid species, mt‐hsp70 has a specific function in kDNA biology, possibly in the processes of kDNA replication, RNA editing or kinetoplast structure.
Encephalitozoon‐Like Organisms in Patients with Alveolar Hydatid Disease: Cell Culture, Ultrastructure, Histoimmunochemical Localization and Seroprevalence
Authors: FURUYA KOJI , SATO CHIAKI , NAGANO HIDEKI , SATO NAOKI , UCHINO JUNCHI
Published: May 2007

ABSTRACT. We found Encephalitozoon‐like organisms in an in vitro culture of a human liver lesion which was due to larval Echinococcus multilocularis. The organisms developed in the same fashion as an Encephalitozoon cunculi. The spores that developed in parasitophorous vacuoles were 2.0–2.6 × 1.1–1.5 μm: each contained a single nucleus and 4–5 polar tubule coils, closely resembling E. cuniculi in its ultrastructure. Mature spores were collected from the supernatants by the use of Percoll centrifugation resulting in the banding of the spores on continuous gradients. We prepared three sorts of spores which were different in buoyant density in 0.15 M NaCl: 1.05–1.07 g/ml spores, 1.12 g/ml spores, and spores of over 1.14 g/ml. Polyclonal antibodies to a pool of each spore preparation were produced in a rabbit and applied to the detection of microsporidian antigen in situ. The histoimmunoperoxidase (HIP) procedure was used to detect the microsporidian antigen in echinococcal liver lesions from patients with alveolar hydatid disease (AHD). Ten echinococcal liver lesions from different AHD patients were examined and four were found to be positive in the HIP test. The Percoll‐separated spores were also used as an antigen to detect for antibodies in the sera from the patients with AHD by Western blotting. Antibodies were detected in 62 (52%) of the 119 AHD patients and in only 8 (5%) of the 159 normal healthy individuals.
Primers Designed for Amplification of Echinococcus multilocularis DNA Amplify the DNA of Encephalitozoon‐Like Spores in the Polymerase Chain Reaction
Authors: FURUYA KOJI , NAGANO HIDEKI , SATO CHIAKI
Published: May 2007

ABSTRACT. Microsporidian spores were developed from cells which were grown in vitro from a human liver lesion which was due to larval Echinococcus multilocularis. The microsporidian spores developed in the same fashion as an Encephalitozoon cuniculi. The Encephalitozoon‐like spores were completely separated on Percoll gradients. The separated spores contained DNA capable of amplification by two different primer sets designed for the polymerase chain reaction (PCR) of E. multilocularis DNA. However, the cell DNA from which microsporidium developed was thoroughly insensitive to the PCR using the E. multilocularis primer sets. The results strongly suggested that Encephalitozoon should be taken into consideration, when DNA isolated from larval E. multilocularis is analyzed.
Intracellular Localization of LRV1 in Infected Leishmania Cells and Epitope Conservation of the Coat Protein
Authors: ZAMORA MIGUEL , SACKS LIISA , STUART KEN
Published: May 2007

ABSTRACT. Polyclonal antibodies were raised against a recombinant fragment of the coat protein of LRV1‐1 to determine the epitope conservation of the coat protein among LRV1 isolates, and the intracellular localization of LRV1 particles in promastigote cells of Leishmania braziliensis. Western blot analysis showed that specific epitopes of the coat protein are highly conserved among isolates from different geographic areas. Using indirect immunofluorescence assays LRV1 viral particles were observed as fluorescent granules, limited to the cytoplasm and with no apparent association to the host organelles or the cell membrane, characteristic of a persistent, non‐infectious virus.
Primary Structure of the Hydrogenosomal Malic Enzyme of Trichomonas vaginalis and Its Relationship to Homologous Enzymes1
Authors: HRDÝ IVAN , MÜLLER MIKLÓS
Published: May 2007

ABSTRACT. The complete nucleotide sequence has been established for two genes (maeA and maeB) coding for different subunits of the hydrogenosomal malic enzyme [malate dehydrogenase (decarboxylating) EC 1.1.1.39] of Trichomonas vaginalis. Two further genes (maeC and maeD) of this enzyme have been partially sequenced. The complete open reading frames code for polypeptides of 567 amino acids in length. These two open reading frames are similar with less than 12 percent pairwise nucleotide differences and less than 9 percent pairwise amino acid differences. The open reading frames of the two partially sequenced genes correspond to the amino‐terminal part of the polypeptides coded and are similar to the corresponding parts of the completely sequenced ones. The deduced translation products of the two complete genes differ in their calculated pI values by 1.5 pH unit. The genes code for polypeptides which contain 12 or 11 amino‐terminal amino‐acyl residues not present in the proteins isolated from the cell. Other hydrogenosomal enzymes also have similar amino‐terminal extensions which probably play a role in organellar targeting and translocation of the newly synthesized polypeptides. A comparison of 19 related enzymes from bacteria and eukaryotes with the maeA product revealed 34–45 percent amino acid identity. Phylogenetic reconstruction based on nonconservative amino acid differences with maximum parsimony (phylogenetic analysis using parsimony, PAUP) and distance based (neighbor‐joining, NJ) methods showed that the T. vaginalis enzyme is the most divergent of all eukaryotic malic enzymes, indicating its long independent evolutionary history.
Lipophosphoglycan Is Present in Distinctly Different Form in Different Entamoeba histolytica Strains and Absent in Entamoeba moshkovskii and Entamoeba invadens1
Authors: SRIVASTAVA GOPAL , ANAND MONIKA TOLA , BHATTACHARYA SUDHA , BHATTACHARYA ALOK
Published: May 2007

ABSTRACT. Lipophosphoglycan has recently been demonstrated on the cell surface of Entamoeba histolytica strain HM‐1:IMSS. A monoclonal antibody against this molecule had failed to react with some other strains of E. histolytica, including the strain Rahman. To determine if a structurally distinct lipophosphoglycan existed in Rahman, [3H]galactose‐labeled glycoconjugates were electrophoresed through sodium dodecyl sulfate polyacrylamide gel electrophoresis. The electrophoretic pattern in Rahman was very different compared to that obtained with strains HM‐1:IMSS and 200:NIH. A number of experiments including sensitivity to mild acid, nitrous acid and phosphoinositol‐specific phospholipase C suggest that the Rahman glycoconjugate is indeed a lipophosphogylcan‐like molecule but distinctly different from that of HM‐1:IMSS. Mild acid‐treated glycoconjugates from Rahman and HM‐1:IMSS revealed the presence of neutral trisaccharides and monosaccharides in Rahman but not in HM‐1:IMSS. Human immune sera from amoebiasis patients and a polyclonal antibody against HM‐1:IMSS liphophosphoglycan both recognized Rahman glycoconjugate. Thus, while lipophosphoglycan molecules from the two strains share common epitopes, they are clearly distinct from each other. Molecules bearing resemblance to lipophosphoglycan could not be detected in other Entamoeba species, namely Entamoeba invadens and Entamoeba moshkovskii.
A Lipoprotein‐Cholesterol‐Albumin Serum Substitute Stimulates Giardia lamblia Encystation Vesicle Formation
Authors: REINER DAVID S. , HETSKO MICHAEL L. , GILLIN FRANCES D.
Published: May 2007

ABSTRACT. We found previously that the A6 clone of Giardia lamblia strain WB that did not encyst in vitro was blocked at an early stage in differentiation, as it did not form encystation secretory vesicles (ESV) efficiently or express cyst antigens, in comparison with the related clone C6. We now report that A6 formed ESV normally in the suckling mouse model. Therefore, we asked whether our serum‐containing encystation media might lack a stimulus or component or contain an inhibitor of ESV formation to which A6 was especially sensitive. We found that replacing bovine serum with a lipoprotein‐cholesterol solution and bovine serum albumin (LPC) in pre‐encystation and encystation media increased ESV formation by both A6 and C6. The % of A6 cells with ESV increased from 8% in BS medium to 48% in LPC medium, compared with 64% and 98% for C6. Similarly, the average number of ESV/positive cell increased from 1.5 in BS medium to 7.7 in LPC medium for A6, and from 13.3 to 19.7 for C6. Moreover, in LPC encystation media, A6 expressed the cyst wall epitope recognized by monoclonal GCSA‐1. Although formation of water‐resistant cysts by A6 was increased >60 fold in LPC media, the numbers of cysts remained only ∼3–15% that of C6. This suggests that LPC may primarily affect early events in encystation and that A6 may require additional factors later in encystation.
Identification of Immunogenic Epitopes of the 170‐kDa Subunit Adhesin of Entamoeba histolytica in Patients with Invasive Amebiasis
Authors: VELAZQUEZ CARLOS , VALETTE IGNEZ , CRUZ MIGUEL , LABRA MARIA‐LUISA , MONTES JULIO , STANLEY SAMUEL L. , CALDERON JESUS
Published: May 2007

ABSTRACT. Entamoeba histolytica causes amebic dysentery (AD) and liver abscess (ALA). Little is known about protective immunity to amebiasis, and studies in this area have been complicated by the paucity of defined ameba antigens. We examined the proliferative responses of peripheral blood mononuclear cells (PBMC) from patients with AD and ALA to a recombinant protein containing a portion of the 170 kDa adhesin of E. histolytica (170CR), and to two synthetic peptides (1 and 2) derived from the 170 kDa sequence that were predicted to contain T cell epitopes. A significant number of patients with AD and ALA had PBMC that proliferated to 170CR molecule, and several individuals with ALA and AD had T cells that recognized one or both peptides. Contrarily, individuals from a non‐endemic region for amebiasis did not respond to 170CR protein, or to both peptides. In regard to antibody response, nine of fifteen patients with ALA showed antibodies to 170CR protein. These same patients had antibodies to peptide 2. We identified peptides from 170‐kDa adhesin that may contain both T and B cell epitopes recognized by some patients with invasive amebiasis. These peptides may be valuable reagents in studies of the immune response to amebiasis.
Trichomonas vaginalis: Ultrastructural Bases of the Cytopathic Effect
Authors: GONZÁLEZ‐ROBLES ARTURO , LÁZARO‐HALLER AMPARO , ESPINOSA‐CANTELLANO MARTHA , ANAYA‐VELÁZQUEZ FERNANDO , MARTÍNEZ‐PALOMO ADOLFO
Published: May 2007

ABSTRACT. The in vitro cytopathic effect of Trichomonas vaginalis on epithelial cells was explored through the interaction of trophozoites of the virulent strain GT‐10 with MDCK monolayers. The interaction was analyzed through electrophysiology, video microscopy, and transmission and scanning electron microscopy. Electrical measurements revealed that living parasites produced severe damage to the cell monolayers within 30 min, manifested as a rapid decrease in transepithelial resistance. Microscopic observations demonstrated that when placed in contact with epithelial cells, trichomonas formed clumps through interdigitations and transient plasma membrane junctions between adjacent parasites. Also, attached trophozoites adopted an ameboid shape. The in vitro cytopathic action of T. vaginalis on MDCK cells was initially evident by modifications of the plasma membrane, resulting in opening of tight junctions, membrane blebbing, and monolayer disruption. After 15 min of interaction the damage was focal, concentrating at sites where parasite clumps adhered to the monolayer. At 30 min practically all MDCK cells were dead, whether or not trichomonas were attached to them. These events were followed by detachment of lysed cells and complete disruption of the monolayer at 60 min. Electron microscopy demonstrated a peculiar form of adhesion that appears to be specific for trichomonas, in which the basal surface of T. vaginalis formed slender channels through which microvilli and cytoplasmic fragments of epithelial cells were internalized. The same sequence of lytic events was found with the less virulent GT‐3 strain. However, the time course of cytolysis with GT‐3 parasites was much slower, and lysis was limited to areas of attachment of T. vaginalis.
Cytoplasm Rescues an Arrhythmic Mutant on the Circadian Rhythm of Mating Reactivity in Paramecium bursaria
Authors: MIWA ISOJI , IZUMO TATSUO , SONODA TOMOKO
Published: May 2007

Cells of an unusual Paramecium bursaria stock (Sj2) expressed rhythmic mating reactivity in a light/dark cycle (LD) and under continuous illumination (LL). When placed in continuous darkness (DD), did not show rhythmicity but rather demonstrated a continuous high mating reactivity. However, mating reactivity was reduced following exposure to a 6‐h light pulse interrupting the DD, and then recovered to its former condition. Genetic analysis showed the arrhythmicity in DD to be a dominant character inherited in a Mendelian ratio. On the other hand, a clone (MCIw) that did not show the rhythmicity in either DD or LL was isolated from the parent stock Sj2w following a 5‐h treatment with 2 μg/ml nitrosoguanidine (MNNG). The MCIw cells expressed weak rhythmicity in LD, but were insensitive to a 6‐h light pulse in DD. The arrhythmicity in LL was inherited cytoplasmically. In addition to this, rhythmicity in LL could be recovered by injection of cytoplasm from the wild‐type cell when the recipient cell was homozygous for the wild‐type nuclear gene (+/+). The cytoplasmic components or factors are assumed to control the functional circadian system and genetically determine the rhythmicity of mating reactivity.
Hypervariability in Gene Copy Number for the Glucose Transporter Genes in Trypanosomes
Authors: BARRETT MICHAEL P. , BRINGAUD FREDERIC , DOUA FELIX , MELVILLE SARA E. , BALTZ THEO
Published: May 2007

The copy number of the gene encoding the carrier protein responsible for the uptake of glucose in bloodstream form Trypanosoma brucei (THT1) was investigated in the genome of 55 different members of the subgenus Trypanozoon. The gene is present in multiple copies in tandem arrays on two homologous chromosomes in these organisms, and copy number varies both intra‐ and interspecifically. Variability is also apparent in the number of genes encoding a second hexose transporter (THT2) which is the only isoform expressed in procyclic organisms. Multiple copies of THT2 are conserved in representatives of the non‐tsetse transmitted species, Trypanosoma evansi and Trypanosoma equiperdum.
Updating the Trachelocercids (Ciliophora, Karyorelictea). I. A Detailed Description of the Infraciliature of Trachelolophos gigas N. G., N. Sp. and T. filum (Dragesco & Dragesco‐Kernéis, 1986) N. Comb
Authors: FOISSNER WILHELM , DRAGESCO JEAN
Published: May 2007

ABSTRACT. Trachelolophos gigas n. g., n. sp. and T. filum (Dragesco & Dragesco‐Kernéis, 1986) n. comb. (basionym: Tracheloraphis filum) were discovered in the mesopsammon of the French Atlantic coast at Roscoff. Their morphology and infraciliature were studied in live and protargol impregnated specimens. The new genus, Trachelolophos, belongs to the family Trachelocercidae and is unique in having a conspicuous ciliary tuft, which is very likely a highly modified brosse, in the oral cavity. The two species investigated have a very similar infraciliature, differing only in morphometric characteristics and in the nuclear configuration. The entire somatic and oral infraciliature consists of dikinetids which have both basal bodies ciliated or only the anterior or posterior ones, depending on the region of the cell. The right side is densely and uniformly ciliated. Its kineties extend onto the left side to the glabrous stripe, where an anterior and posterior secant system are formed, reducing the number of kineties in the narrowed neck and tail region. The left side bears a narrow glabrous stripe bordered by slightly irregularly arranged dikinetids having rather stiff cilia (bristles), possibly forming an uninterrupted, prolate ellipsoidal (bristle) kinety as indicated by their ciliation. The bristle kinety commences subapically at the right margin of the glabrous stripe, extends posteriorly, then anteriorly at the left, to end up at the right margin again. The dikinetids of the right posterior portion of the bristle kinety have the posterior basal bodies ciliated, whereas the anterior basal bodies are ciliated in its left and right anterior portion. The ends of the bristle kinety meet distinctly subapically at the right margin of the glabrous stripe, as indicated by the diametrically opposed ciliation of the dikinetids. The anterior region (head) of the cell bears a distinct circumoral kinety composed of very regularly arranged dikinetids, associated with nematodesmata forming an oral basket together with the nematodesmal bundles originating from the oralized somatic dikinetids at the anterior end of the somatic kineties. The systematics of trachelocercid ciliates are briefly reviewed and discussed.
Identification of a Microsporidian Polar Tube Protein Reactive Monoclonal Antibody
Authors: KEOHANE ELAINE M. , TAKVORIAN PETER M. , CALI ANN , TANOWITZ HERBERT B. , WITTNER MURRAY , WEISS LOUIS M.
Published: May 2007

ABSTRACT. The microsporidia are characterized by spores containing a single polar tube that coils around the sporoplasm. When triggered by appropriate stimuli, the polar tube rapidly discharges out of the spore forming a hollow tube. The sporoplasm passes out of the spore through this tube serving as a unique vehicle of infection. Due to the unusual functional and solubility properties of the polar tube, the proteins comprising it are likely to be members of a protein family with a highly conserved amino acid composition among the various microsporidia. Polar tube proteins were separated from the majority of other proteins in glass bead disrupted spores of Glugea americanus using sequential 1% sodium dodecyl sulfate (SDS) and 9M urea extractions. The resultant spore pellet demonstrated broken, empty spore coats and numerous polar tubes in straight and twisted formations by negative stain transmission electron microscopy. After subsequent incubation of the pellet with 2% dithiothreitol (DTT), empty spore coats were still observed but the polar tubes were no longer present in the pellet. The DTT supernatant demonstrated four major protein bands by SDS‐PAGE: 23, 27, 34 and 43 kDa. Monoclonal antibodies were produced to these proteins using Hunter's Titermax adjuvant. Mab 3C8.23.1 which cross‐reacted with a 43‐kDa antigen by immunoblot analyis, demonstrated strong reactivity with the polar tube of G. americanus spores by immunogold electron microscopy. This antibody will be useful in further characterization of polar tube proteins and may lead to novel diagnostic and therapeutic reagents.
In Vitro Excystation of Spironucleus muris
Authors: KOUDELA BŘETISLAV , KARR CRAIG D. , MEYER ERNEST A. , MEYER BARBARA , JARROLL EDWARD L.
Published: May 2007

ABSTRACT. In vitro excystation of Spironucleus muris cysts, purified by sequential sucrose and Percoll gradients from mouse feces, was studied. Three in vitro excystation procedures, used for Giardia, were assessed to determine the most useful method. Excystation was monitored by light microscopy and subsequently characterized by transmission and scanning electron microscopy. Spironucleus muris excysted routinely at a level greater than 90% when induced in Hanks’ balanced salt solution containing sodium bicarbonate at pH 2.0 and transferred to Tyrodes’ salt solution as an excystation medium. Similarly, high rates of excystation were recorded after induction of S. muris cysts in 0.1 M potassium phosphate buffer (pH 7.0) with sodium bicarbonate and excystation in trypticase‐yeast extract‐iron medium (TYI medium) or phosphate‐buffered saline. A lower rate and percentage of excystation were observed after induction of S. muris cysts in an aqueous hydrochloric acid solution (pH 2.0) followed by excystation in TYI medium. All excystation methods produced extremely active S. muris trophozoites with normal morphology. Nonexcysting S. muris cysts have a wall composed of an outer fibrous and an inner membranous portion. Following induction, numerous vesicles appeared in the peritrophic space. Excystation began by the cyst wall opening at one pole, and the anterior part of the trophozoite protruding from the cyst wall. The trophozoite emerged progressively from the cyst wall and the empty cyst wall appeared to collapse. Excysted trophozoites exhibited normal morphological features of S. muris trophozoites isolated from the mouse intestine.
Phagocytosis in Dictyostelium: Nibbling, Eating and Cannibalism
Authors: LEWIS KEITH E. , O'DAY DANTON H.
Published: May 2007

ABSTRACT. Phagocytosis is a highly conserved biological process that serves numerous functions in a wide variety of organisms. Over the past few decades Dictyostelium has proven to be an excellent organism for investigations in cell biology and this is certainly no less the case for a study of phagocytosis. This review examines three distinct phagocytic activities which have been characterized in Dictyostelium. The first, “vegetative phagocytosis,” represents the classical eukaryotic microbial uptake of food particles (bacteria). The second, a predatory form of phagocytosis, arises when one species such as Dictyostelium caveatum attacks another species of slime mold, engulfing small pieces of the target prey. This has been termed “cell nibbling.” The third phagocytic process is “sexual cannibalistic phagocytosis.” In this situation a zygote giant cell, having arisen from the fusion of gametic amoebae, attracts unfused nonzygotic amoebae of the same species and engulfs them as a food source. While cell nibbling has not been actively studied, vegetative and sexual cannibalistic phagocytosis have received varying amounts of attention leading to the idea that some of the elements (e.g., glycoprotein receptors and a Gαs subunit) involved in certain of these phagocytic events may be the same. On the other hand, some unique events (e.g., filopodial induction in prey by D. caveatum) are also worthy of further investigation. Among other things, the presence of self‐nonself recognition, the existence of opsonin‐like substances and the presence of signal transduction elements (e.g., an A2‐like receptor that negatively modulates sexual phagocytosis) once considered to be extant only in higher organisms suggest that much can be learned about phagocytosis in general by further studies in the classic, eukaryotic microbe Dictyostelium discoideum and related species.
A New Family of Entodiniomorph Protozoa from the Marsupial Forestomach, with Descriptions of a New Genus and Five New Species
Authors: DEHORITY BURK A.
Published: May 2007

ABSTRACT. A unique group of entodiniomorph protozoa was found in forestomach contents from quokka (Setonix brachyurus), western grey kangaroo (Macropus fuliginosus), red kangaroo (Macropus rufus) and euro (Macropus robustus erubescens). A new genus, Macropodinium n.g., containing five new species, is described. Three species are described from forestomach contents of the quokka: Macropodinium baldense n. sp., Macropodinium moiri n. sp. and Macropodinium setonixum n. sp. A single species, Macropodinium ennuensis n. sp., is described from the red kangaroo and euro. The last species, Macropodinium yalanbense n. sp., is described in forestomach contents from the western grey kangaroo. At least three distinct features in the new genus are incompatible with any of the described families in the order Entodiniomorphida. On this basis, the new family Macropodiniidae has been created.
Commitment to Division in Ciliate Cell Cycles
Authors: ADL SINA M. , BERGER JAMES D.
Published: May 2007

Near the end of the cell cycle, ciliates commit irreversibly to cell division. The point of commitment occurs at the time of oral polykinetid assembly and micronuclear anaphase. The commitment is a checkpoint which requisites a threshold cell mass/ DNA ratio and stomatogenesis. It is also a physiological transition point, involving cdk protein kinases similar to those of other eukaryotes. Both P34 kD and P36 kD kinases, similar to the S. pombe cdc2 kinases, have been described to have activity as monomers. Subsequent to commitment to division, dramatic cytoskeletal modifications occur for separation of organelles, cortex morphogenesis and cytokinesis. Numerous mutants affecting cytoskeletal function associated with the division process have been obtained in several species. Of these, only the ccl mutant in Paramecium affects cell cycle progression prior to commitment to division. The material reviewed is used to speculate about the mechanisms of regulation of pre‐fission morphogenesis and cell division related processes in ciliates.
Stomatogenesis in Kyaroikeus cetarius (Dysteriina, Kyaroikeidae, N. Fam.): Clues to Its Systematic Placement
Authors: SNIEZEK JAMES H. , COATS D. WAYNE
Published: May 2007

Morphogenesis in Kyaroikeus cetarius, a large phyllopharyngean parasite of odontocete Cetacea, begins with the midventral proliferation of four short rows of kinetosomes immediately to the left of four pre‐existing, somatic kinetofragments. The resulting field of eight short kineties is located within a shallow depression that gradually elongates and deepens to form the oral cavity of the opisthe. The four right‐most of these kineties sink into the developing oral cavity and divide transversely, producing three distinct sets of kinetofragments. The anterior and posterior sets each consist of four small fragments and ultimately produce the mid‐ventral, somatic kinetofragments of the proter and the opisthe, respectively. In addition, some kinetosomes from the anterior set give rise to the secretory organelle complex of the proter. The central group consists of two long and one short kinetofragment, which represent anlagen for the opisthe circumoral and preoral kineties, respectively. These anlagen migrate progressively anteriad while undergoing a pronounced counterclockwise rotation that eventually inverts the developing oral kineties. Simultaneously, two broad sheets of microtubules, one beneath each circumoral anlage, extend deep into the cytoplasm and unite to line the cytopharynx of the opisthe. Meanwhile, the remainder of the eight original kinetofragments move laterally out of the ventral depression to produce the left ciliary field of the opisthe. Morphogenetic events observed in K. cetarius are typical of cyrtophorid Phyllopharyngia and support inclusion of this genus within suborder Dysteriina.
Ca2+‐Dependence of Conoid Extrusion in Toxoplasma gondii Tachyzoites
Authors: MONDRAGON RICARDO , FRIXIONE EUGENIO
Published: May 2007

The role of Ca2+ in conoid extrusion was investigated in isolated Toxoplasma gondii tachyzoites by treatment with Ca2+‐ionophores, Ca2+‐chelating agents and an inhibitor of the Ca2+‐ATPase at the endoplasmic reticulum. The results were evaluated by light phase‐contrast microscopy and electron microscopy. lonomycin (0.5‐1 μM) caused an immediate and sustained extrusion of the conoid in up to 80% of the tachyzoites, depending on the concentrations of ionophore and Ca2+ in the medium. However, over 50% of the tachyzoites extruded the conoid when treated with ionomycin in Ca2+‐free saline complemented with EGTA. The effect of ionomycin was reversible and could be induced a second time in about half of the responsive population. Similar results were obtained with A23187. Conoid extrusion induced by ionomycin in Ca2+‐free medium was almost completely abolished when the tachyzoites were previously loaded with a permeable compound known to chelate intracellular Ca2+ (BAPTA/AM; 25μM). On the other hand, exposure of tachyzoites to the Ca2+‐ATPase inhibitor thapsigargin (0.5‐1μM) produced significant extrusion of the conoid. Tachyzoites loaded with BAPTA/AM as well as those treated with ionomycin, i.e. with conoids paralyzed in opposite positions, had a diminished capacity to invade cultured epithelial cells. A substantial reduction in the response to stimulation by ionomycin was found also in parasites treated with cytochalasin‐D, a drug that depolymerizes actin‐filaments. The results suggest that Ca2+‐release from internal stores may act as a key signal to activate a mechanism of conoid extrusion probably mediated, at least in part, by actin‐filaments.
Epigenetic Regulation of Programmed Genomic Rearrangements in Paramecium aurelia1
Authors: MEYER ERIC , DUHARCOURT S.
Published: May 2007

ABSTRACT In ciliates, development of the polyploid somatic macronucleus after sexual events involves extensive and reproducible rearrangements of the germ‐line genome, including chromosome fragmentation and precise excision of numerous internal sequence elements. In Paramecium aurelia, alternative macronuclear versions of the same germ‐line genome can be maternally inherited across sexual generations, showing that rearrangement patterns are not strictly determined by the germ‐line sequence. Homology‐dependent maternal effects can be evidenced by transformation of the vegetative macronucleus with cloned macronuclear sequences: new fragmentation patterns or internal deletions are specifically induced during differentiation of a new macronucleus, in sexual progeny of transformed clones. Furthermore, transformation of the maternal macronucleus with germ‐line sequences containing internal eliminated sequences (short single‐copy elements) can result in a specific inhibition of the excision of the same elements in the zygotic macronucleus. These experiments show that the processing of many germ‐line sequences in the developing macronucleus is sensitive to the structure and copy number of homologous sequences in the maternal macronucleus. The generality and sequence specificity of this trans‐nuclear, epigenetic regulation of rearrangements suggest that it is mediated by pairing interactions between germ‐line sequences and sequences imported from the maternal macronucleus.