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Demonstration of glycosomes (microbodies) in the Bodonid flagellate Trypanoplasma borelli (Protozoa, Kinetoplastida).

Homogenates of Trypanoplasma borelli were subjected to subcellular fractionation by sequential differential and isopycnic centrifugation in sucrose. Glycerol-3-phosphate dehydrogenase and the glycolytic enzymes, glucosephosphate isomerase and triosephosphate isomerase, as well as the peroxisomal marker enzyme catalase were mainly, or in part, associated with sedimentable particles that had a buoyant density in sucrose of 1.22 g cm-3. Moreover, triosephosphate isomerase exhibited latency, both in total homogenates and in the particulate fraction. Electron microscopy of thin sections of T. borelli revealed the presence of microbodies that gave a positive reaction for catalase. Pyruvate kinase behaved as a typical soluble enzyme. It was stimulated by micromolar concentrations of fructose 2,6-bisphosphate and this stimulation was counteracted by inorganic phosphate in the millimolar range. The enzymes involved in the synthesis and degradation of fructose 2,6-bisphosphate, 6-phosphofructo-2-kinase and fructose-2,6-bisphosphatase, were both present in T. borelli and behaved as soluble enzymes. We conclude that in T. borelli the glycolytic pathway is compartmentalized in a way similar to that found in another Kinetoplastid family, the Trypanosomatidae, where seven glycolytic enzymes and two enzymes of glycerol metabolism are associated with glycosomes. Apparently the presence of glycosomes is a characteristic of all Kinetoplastida.

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Phylogeny of trypanosomes as inferred from the small and large subunit rRNAs: implications for the evolution of parasitism in the trypanosomatid protozoa.

Sequences of the small rRNA genes and partial sequences of the large rRNA genes were obtained by PCR amplification from a variety of vertebrate trypanosomes. The trypanosome species and hosts included Trypanosoma avium from a bird, T. rotatorium from an amphibian, T. boissoni from an elasmobranch, T. triglae from a marine teleost and T. carassii from a freshwater teleost. Phylogenetic relationships among these species and other representatives of the family Trypanosomatidae were inferred using maximum likelihood, maximum parsimony and evolutionary parsimony. The trypanosomatid tree was rooted using rRNA sequences from two species from the suborder Bodonina. All methods showed that the mammalian parasite, Trypanosoma brucei, constitutes the earliest divergent branch. The remaining trypanosomes formed a monophyletic group. Within this group, the bird trypanosome was grouped with T. cruzi, while the elasmobranch trypanosome and the two fish trypanosome species formed a group with an affinity to T. rotatorium. Our results provide no evidence for co-evolution of trypanosomatids and their hosts, either vertebrate or invertebrate. This suggests that evolution of trypanosomatids was accompanied by secondary acquisitions of hosts and habitats.

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Microtubular cytoskeletons of parasitic protozoa.

Shape, motility and division of eukaryotic cells are all determined to some extent by intracellular microtubules. Parasites, particularly protozoan parasites, offer important models for the study of microtubule organization, which may also provide useful leads for novel chemotherapeutic agents against parasite-specific forms of these organelles. Research on the biology of microtubules has advanced rapidly in the last 2-3 years, from purely descriptive studies to on understanding of some of the genetic and biochemical aspects. In this article, David Russell and Jean-François Dubremetz summarize the progress and discuss the potential of tubulin as a drug target for antiprotozool chemotherapy.

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Site-specific retrotransposons of the trypanosomatid protozoa.

Retrotransposons are mobile genetic elements that have invaded a wide variety of organisms. While these mobile elements share gene homologies and structural features with retroviruses, they have lost the ability to produce infectious particles. Typically these elements are 5-10 kilobases (kb) in length, are conserved in their structural organization and are present in many copies in the genomes into which they have integrated(1). Retrotransposons generally interrupt their host genome promiscuously and thus cause a variety of random effects. In general, their insertion results in mutations, inversions, deletions or rearrangements among host sequences. All of these changes are thought to add to the plasticity of the host genome and thus contribute to a faster pace of evolutionary development. However, because of the random nature of insertions, it has been difficult to attribute any one specific function to these diverse elements. Here, Serap Aksoy describes a newly recognized family of mobile elements that are different from most retrotransposons in that they have the ability to integrate into specific host sequences.

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The worm and the protozoa: stereotyped responses or distinct antigens?

Studies of parasitic diseases have provided the best in vivo correlates of the division of CD4+ helper T cells into distinct functional phenotypes, designated T(H)I and T(H)2, that mediate the balanced regulation of cellular and humoral immunity. In this article, Steven Reiner and Richard Locksley focus on why parasitic infections tend to generate such clearly polarized responses and emphasize that early events that mediate maturation signals towards T(H)1- or T(H)2-effector and memory cells remain incompletely defined. Effective vaccination that seeks to mold the developing immune response will need to consider the role of interleukins and various cell-surface molecules that have been identified, thus far, to influence CD4 subset differentiation.

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Protozoa as model systems for the study of cellular responses to altered gravity conditions.

The orientation behavior of Paramecium changed in a similar way after transition to conditions of free-fall in a sounding rocket and after transition to conditions of simulated weightlessness on a fast rotating clinostat. After a period of residual orientation, Paramecium cells distributed themselves randomly 80 s (120 s) after onset of free-fall (simulated weightlessness). Swimming velocity increased significantly; however, the increase was transient and subsided after 3 min in the rocket experiments, while the velocity remained enhanced even during 2 h of rotation on a fast clinostat. Trichocysts were present and without morphological changes in Paramecium cells which had been exposed to a rocket flight, as well as to fast or slow rotation on a clinostat. Regeneration of the oral apparatus of Stentor and morphogenesis of Eufolliculina proceeded normally on the clinostat. The results demonstrate that the clinostat is a useful tool to simulate the conditions of weightlessness on earth and to detect gravisensitive cellular functions.

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Application of ATP measurement to evaluation of the growth of parasitic protozoa in vitro with a special reference to Pneumocystis carinii.

1. There was a significant correlation between the increase in the number of Entamoeba histolytica, Trichomonas vaginalis, Giardia lamblia and Leishmania donovani in culture, and their ATP contents determined by luciferase reaction. 2. The similar correlation was also demonstrated between the decreased number of E. histolytica in the presence of an anti-amebic quassinoid and the nucleotide content in vitro. 3. In the case of Pneumocystis carinii, the numbers of the organism remained relatively constant in culture for at least 7 days without growth; however, the ATP content dropped rapidly in 1 to 3 days except in RPMI 1640. 4. The possibility that ATP determination of P. carinii is complicated by the host cell nucleotide seemed to be excluded, since the concentration of this nucleotide in normal lung was almost negligible. These observations suggest that the present procedure is useful for evaluating the growth and viability of these organisms in vitro.

Adenosine Triphosphate↗

The myoneme of the Acantharia (Protozoa): A new model of cellular motility.

The myonemes of Acantharia are made of bundles of microfilaments twisted up in elementary microstrands of two. Myonemes exhibit three kinds of movements: slow contraction accompanied by undulations of the organelle, quick contraction and subsequent relaxation. In vivo, contraction-relaxation cycles were studied with normal, time-lapse and high speed film-recordings (1000 frames/s). Kinetic parameters of these movements are given. All the movements are actively produced by the organelle itself and are dependant on calcium concentration. An hypothesis about the way myoneme microfilaments might interact to generate motile forces is given. The most closely related system might be the myoneme of the dinoflagellate Noctiluca miliaris.

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The evolution of RNA editing in kinetoplastid protozoa.

RNA editing alters messenger RNA transcripts by the addition and deletion of uridine residues. This remarkable process corrects frameshift mutations and replaces AUG initiation codons in most trypanosomatid mitochondrial RNA transcripts. Sometimes, however, RNA editing results in open reading frames whose sequences differ by greater than 50% from the original RNA sequences. In this paper, we will explore two models for the evolution of RNA editing in kinetoplastid protoza.

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Merogony and gametogony of Eimeria mccordocki (Protozoa-Eimeriidae) in the mule deer, Odocoileus h. hemionus.

Eimeria mccordocki and E. madisonensis oocysts were isolated from feces of 21 of 40 captive mule deer in Fort Collins, Colorado. The two species were separated from each other by infecting one mule deer fawn, and the life cycle of E. mccordocki was studied for the first time. Four to six-weeks-old mule deer fawns were inoculated orally with E. mccordocki and killed 9, 13 and 15 days after infection. Asexual and sexual stages of life cycle developed in the ileum of mule deer, only in the surface epithelial cells of the villi. The asexual stages consisted of two generations of meronts.

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A review on vaccination against protozoa and arthropods of veterinary importance.

The recent advances in immunology and biotechnology have stimulated much research on the control of parasitic diseases through vaccination. This is a review of the state of the art regarding important protozoan and arthropod veterinary parasites. A live oocyst vaccine for avian coccidiosis is still in use but much work has been done on the identification, cloning, and assay of protective antigens. The sporozoites of Eimeria tenella have been the preferred subject and at least four recombinant antigens have already been tested with partial success. Premunization against babesiosis is still widely used in Latin America as is a live vaccine with attenuated parasites in Australia. At least three Babesia bovis and three Babesia bigemina antigens that generate partial protection have been produced as recombinant proteins. A vaccine against canine babesiosis is being commercialized in France. Infection-treatment is still used to vaccinate against Theileria parva and a schizont vaccine against Theileria annulata. Recombinant sporozoite antigens have been assayed with partial success against both species but the identification and administration of protective schizont antigens, regarded as the most important, still requires considerable work. The immunological control of African trypanosomoses is still impaired by the antigenic variation that the parasites experience during the infection. Although some possibilities exist, most specialists are pessimistic about the promise of developing a vaccine in the near future. Control of Boophilus ticks with an occult tick intestine recombinant antigen seems to have potential in inhibiting reproduction of the tick but salivary antigens appear to be more effective at inhibiting feeding and pathogen transmission. Vaccination with a Hypoderma protein, recently cloned, has induced 90% protection against subsequent infestations. It is very likely that effective vaccines against veterinary parasites will become available in the near future.

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