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ABSTRACT: Differentiation into infectious cysts (encystation) and multiplication of pathogenic trophozoites after hatching from the cyst (excystation) are fundamental processes in the life cycle of the human intestinal parasite Giardia intestinalis. During encystation, a bi-nucleated trophozoite transforms to a dormant tetra-nucleated cyst enveloped by a protective cyst wall. Nuclear division during encystation is not followed by cytokinesis. In contrast to the well-studied mechanism of cyst wall formation, information on nuclei behavior is incomplete and basic cytological data are lacking. Here we present evidence that (1) the nuclei divide by semi-open mitosis during early encystment; (2) the daughter nuclei coming from different parent nuclei are always arranged in pairs; (3) in both pairs, the nuclei are interconnected via bridges formed by fusion of their nuclear envelopes; (4) each interconnected nuclear pair is associated with one basal body tetrad of the undivided diplomonad mastigont; and (5) the interconnection between nuclei persists through the cyst stage being a characteristic feature of encysted Giardia. Based on the presented results, a model of nuclei behavior during Giardia differentiation is proposed.
Protist 12/2011; 163(3):465-79. · 3.14 Impact Factor
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ABSTRACT: Differentiation into highly resistant double-walled cysts is a major mechanism allowing amphizoic acanthamoebae to survive under long-lasting, unfavourable environmental conditions. We found that relatively low concentrations of methanol, acetone or DMSO stimulate promptAcanthamoebadifferentiation into a rounded cyst-like stage with a single envelope. To address whether this rapid response differs from the encystment, time-dependent changes in cell surface characteristics and cyst-specific gene expression were monitored in encystating cells and cells differentiating under methanol treatment using microscopic, lectin-binding, PCR and resistance studies. In contrast to the encystment: (1) a single-layered amorphous mannose/glucose coat was the only envelope assembled on the surface of the solvent-treated cells, (2) the cyst-specific protein (CSP21) was not expressed, (3) the coat did not protect cells against acidic pH and (4) in solvent-free encystment medium, the coated cells did not assemble the double-layered wall, thus indicating that these cells were not immature cysts. These findings lead us to specify a terminal stage of rapidAcanthamoebadifferentiation elicited by acute organic solvent stress as "pseudocyst", and to suggest that encystation and pseudocyst formation are distinct stress responses. Moreover, the possibility exists that pseudocysts might form in response to certain contact lens solutions thus increasing resistance of acanthamoebae to disinfecting agents.
Protist 01/2011; 162(1):58-69. · 3.14 Impact Factor
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ABSTRACT: Propylene glycol used as an ophthalmic demulcent in certain contact-lens care systems has been included recently among factors responsible for increasing Acanthamoeba keratitis. In this study, we provide evidence that propylene glycol as well as examined contact-lens solutions containing it induce rapid differentiation of acanthamoebae into pseudocysts. The partial resistance of the pseudocysts and their reversibility to viable trophozoites even after 24-h exposure to the contact-lens solutions indicate a potential risk of infection to contact-lens users.
Experimental Parasitology 01/2011; 127(1):326-8. · 2.12 Impact Factor
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ABSTRACT: We propose a new classification of Parabasalia which is congruent with both ultrastructural and molecular-phylogenetic studies. We identify six main parabasalid lineages and give them the rank of class: Hypotrichomonadea, Trichomonadea, Tritrichomonadea, Cristamonadea, Trichonymphea, and Spirotrichonymphea. Trichomonadea is characterized by a single mastigont and by the absence of both a comb-like structure and an infrakinetosomal body. Most representatives also possess a lamelliform undulating membrane. Trichomonadea is divided into two monophyletic orders, Trichomonadida (family Trichomonadidae; with a B-type costa) and Honigbergiellida (families Honigbergiellidae, Hexamastigidae and Tricercomitidae; without a costa). The class Tritrichomonadea, with a single order Tritrichomonadida, is ancestrally characterized by a single mastigont with four flagella, and both a comb-like structure and an infrakinetosomal body. The morphologically most complex representatives (family Tritrichomonadidae) possess in addition a rail-type undulating membrane, an A-type costa, and a suprakinetosomal body. These last three characters are absent in families Monocercomonadidae and Simplicimonadidae. The remaining tritrichomonadids, Dientamoebidae, have undergone reductive evolution. Cristamonads (Cristamonadea) are morphologically derived from tritrichomonads. Because we are unable to determine morphologically homogenous monophyletic lineages within cristamonads, we classify all cristamonads into a single family, Lophomonadidae. Hypotrichomonadea, comprising the genera Trichomitus and Hypotrichomonas, resembles Tritrichomonadea by an A-type costa, and by the presence of a comb-like structure in the mastigont. However, they do not possess an infrakinetosomal body, and are not specifically related to Tritrichomonadea in molecular-phylogenetic analyses. Moreover, unlike Tritrichomonadea, Hypotrichomonadea possesses a lamelliform undulating membrane. The remaining parabasalids are of complex morphology and belong to the classes Trichonymphea and Spirotrichonymphea. A new parabasalid genus, Simplicimonas (Tritrichomonadea), and three new species, Tetratrichomonas undula, Hexamastix coercens and Simplicimonas similis, are described.
Protist 07/2010; 161(3):400-33. · 3.14 Impact Factor
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03/2010; , ISBN: 9780470688618
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Molecular Phylogenetics and Evolution 06/2008; 48(2):770-5. · 3.61 Impact Factor
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ABSTRACT: Fornicata is a relatively recently established group of protists that includes the diplokaryotic diplomonads (which have two similar nuclei per cell), and the monokaryotic enteromonads, retortamonads and Carpediemonas, with the more typical one nucleus per cell. The monophyly of the group was confirmed by molecular phylogenetic studies, but neither the internal phylogeny nor its position on the eukaryotic tree has been clearly resolved.
Here we have introduced data for three genes (SSU rRNA, alpha-tubulin and HSP90) with a wide taxonomic sampling of Fornicata, including ten isolates of enteromonads, representing the genera Trimitus and Enteromonas, and a new undescribed enteromonad genus. The diplomonad sequences formed two main clades in individual gene and combined gene analyses, with Giardia (and Octomitus) on one side of the basal divergence and Spironucleus, Hexamita and Trepomonas on the other. Contrary to earlier evolutionary scenarios, none of the studied enteromonads appeared basal to diplokaryotic diplomonads. Instead, the enteromonad isolates were all robustly situated within the second of the two diplomonad clades. Furthermore, our analyses suggested that enteromonads do not constitute a monophyletic group, and enteromonad monophyly was statistically rejected in 'approximately unbiased' tests of the combined gene data.
We suggest that all higher taxa intended to unite multiple enteromonad genera be abandoned, that Trimitus and Enteromonas be considered as part of Hexamitinae, and that the term 'enteromonads' be used in a strictly utilitarian sense. Our result suggests either that the diplokaryotic condition characteristic of diplomonads arose several times independently, or that the monokaryotic cell of enteromonads originated several times independently by secondary reduction from the diplokaryotic state. Both scenarios are evolutionarily complex. More comparative data on the similarity of the genomes of the two nuclei of diplomonads will be necessary to resolve which evolutionary scenario is more probable.
BMC Evolutionary Biology 01/2008; 8:205. · 3.52 Impact Factor
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ABSTRACT: Abstract
Background
Fornicata is a relatively recently established group of protists that includes the diplokaryotic diplomonads (which have two similar nuclei per cell), and the monokaryotic enteromonads, retortamonads and Carpediemonas , with the more typical one nucleus per cell. The monophyly of the group was confirmed by molecular phylogenetic studies, but neither the internal phylogeny nor its position on the eukaryotic tree has been clearly resolved.
Results
Here we have introduced data for three genes (SSU rRNA, α-tubulin and HSP90) with a wide taxonomic sampling of Fornicata, including ten isolates of enteromonads, representing the genera Trimitus and Enteromonas , and a new undescribed enteromonad genus. The diplomonad sequences formed two main clades in individual gene and combined gene analyses, with Giardia (and Octomitus ) on one side of the basal divergence and Spironucleus , Hexamita and Trepomonas on the other. Contrary to earlier evolutionary scenarios, none of the studied enteromonads appeared basal to diplokaryotic diplomonads. Instead, the enteromonad isolates were all robustly situated within the second of the two diplomonad clades. Furthermore, our analyses suggested that enteromonads do not constitute a monophyletic group, and enteromonad monophyly was statistically rejected in 'approximately unbiased' tests of the combined gene data.
Conclusion
We suggest that all higher taxa intended to unite multiple enteromonad genera be abandoned, that Trimitus and Enteromonas be considered as part of Hexamitinae, and that the term 'enteromonads' be used in a strictly utilitarian sense. Our result suggests either that the diplokaryotic condition characteristic of diplomonads arose several times independently, or that the monokaryotic cell of enteromonads originated several times independently by secondary reduction from the diplokaryotic state. Both scenarios are evolutionarily complex. More comparative data on the similarity of the genomes of the two nuclei of diplomonads will be necessary to resolve which evolutionary scenario is more probable.
BMC Evolutionary Biology. 01/2008;
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ABSTRACT: Derivatives of 5-nitroimidazole, such as metronidazole or tinidazole, are the drugs of choice for
treatment of sexually transmitted infections of humans caused by the parasitic protist Trichomonas
vaginalis. These drugs with selective activity against anaerobic and microaerophilic microorganisms
enter the trichomonad cell and accumulate in hydrogenosomes, where their antimicrobial properties are activated.
In this chapter we discuss metabolic pathways of hydrogenosomes involved in metronidazole activation. We
also summarize present knowledge on the development and biochemical mechanisms of metronidazole resistance
in T.vaginalis and the related cattle parasite Tritrichomonas
foetus. Implications of data from the T.vaginalis genome
project suggesting the presence of novel mechanisms of drug resistance are also considered.
12/2007: pages 179-199;
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ABSTRACT: The family Monocercomonadidae (Parabasala, Trichomonadida) is characterized by the absence of a costa and in most species also of an undulating membrane; both of which are typical structures of trichomonadids. We have examined 25 isolates of Monocercomonadidae species by sequencing of the SSU rDNA and the ITS region and by light and transmission electron microscopy. The isolates formed three distinct phylogenetically unrelated clades: (1) Monocercomonas colubrorum, (2) Monocercomonas ruminantium together with a strain ATCC 50321 designated as Pseudotrichomonas keilini, and (3) Hexamastix. Twenty isolates of Monocercomonas colubrorum split into three clades with no host-specificity. The morphological differences among clades were insufficient to classify them as a separate species. Non-monophyly of the cattle commensal Monocercomonas ruminantium with the type species Monocercomonas colubrorum and absence of Pseudotrichomonas characters in the free-living strain ATCC 50321 led to their reclassification into a new genus (Honigbergiella gen. nov.). The close relationship of these strains indicates a recent switch between a free-living habit and endobiosis. Two strains of Hexamastix represented different species -Hexamastix kirbyi Honigberg 1955 and Hexamastix mitis sp. nov. Polyphyly of the Monocercomonadidae confirmed that the absence of a costa and an undulating membrane are not taxonomically significant characters and were probably secondarily lost in some or all clades. Our observations, however, indicated that other characters - infrakinetosomal body, comb-like structure, marginal lamella, and the type of axostyle - are fully consistent with the position of Monocercomonadidae species in the parabasalian tree and are, therefore, reasonable taxonomic characters.
Protist 08/2007; 158(3):365-83. · 3.14 Impact Factor
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ABSTRACT: The phylogeny and unexpected distribution of hydrogenase-related genes among extant organisms suggest an ancient and crucial
role for members of this gene family in eukaryotes and are consistent with hydrogen syntrophy based models of eukaryogenesis.
For the protists studied, the available data are consistent with a mitochondrial origin for the hydrogenosomes. But one needs
to realize that the origin of the enzymes need not necessarily be the same as the origin of the organellar structure. The
hydrogenosomes may have originated several times in the course of evolution, with lateral transfer of genes from other eukaryotes
or anaerobic bacteria. At present, the lack of a well-supported sister group to the eukaryotes prevents discrimination between
the ideas presented in the hydrogen and syntrophic hypotheses. Furthermore, the most parsimonious explanation of these observations
is that many eukaryotes, such as the green algae, retained ancestral energy metabolic genes that were available for recruitment
during adaptation to anaerobic and, in at least one case, photosynthetic lifestyles, a remarkable demonstration of the plasticity
of eukaryote molecular physiology.
05/2007: pages 113-127;
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ABSTRACT: The random amplified polymorphic DNA (RAPD) technique was used for phylogenetic analysis of trichomonads, for intraspecies genealogical study of Trichomonas vaginalis strains, and for assessment of intrastrain polymorphism in Trichomonas vaginalis. The phylogenetic tree for 12 trichomonad species showed certain discrepancies with current models of trichomonad evolution. However, it shows that RAPD traits retain phylogenetically relevant information. The results of intraspecies analyses of 18 Trichomonas vaginalis strains suggested some concordance between the genetic relationship of strains and their geographic origin. They also suggested a concordance between the strain genetic relationships and the resistance to metronidazole. A concordance was also found with respect to the severity of disease observed in donor patients but not with the results of laboratory virulence assays. No concordance was found between genetic relationship of strains and strain infection with a dsRNA Trichomonas vaginalis virus (TVV). The latter suggests that TVV might be transmitted horizontally among Trichomonas vaginalis populations. The identity of RAPD patterns of clones isolated from in vitro cultures and those of the cultures reisolated independently from the same patient within a period of six weeks suggests that individual Trichomonas vaginalis strains are not polymorphic and that the RAPD patterns are stable. Therefore, the RAPD technique seems useful for addressing various clinically relevant issues.
Journal of Eukaryotic Microbiology 05/2007; 44(6):545 - 552. · 2.66 Impact Factor
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ABSTRACT: Trophozoites of Giardia are equipped with a special organelle of attachment, essential for parasite survival and pathogenicity, the ventral disc. Although its basic structure is well established, its reorganization and assembly during cell replication is poorly understood. We addressed some of these problems with aid of conventional, confocal and electron microscopy. We found that dividing Giardia alternates attached and free swimming phases in accordance with functional competence of the parent or newly assembled discs. The division started in attached cells by detachment of the disc microtubules from basal bodies. Shortening and eventual loss of the giardin microribbons, and unfolding of the microtubular layer resulting in collapse of the disc chamber and parasite detachment underlined gradual disassembly of the parent disc skeleton. Two daughter discs assembled on the dorsal side of the attached cell, with their ventral sides exposed on the parent cell surface and their microtubular skeletons growing in counter-clockwise direction. A depression between the assembling discs marked the cleavage plane. The splitting continued during the free-swimming phase with ventral-ventral axial symmetry in a plane of the daughter discs. Finally, the daughter cells with fully developed discs but still connected tail to tail by a cytoplasmic bridge, attached to a substrate and terminated the division by a process resembling adhesion-dependent cytokinesis. The mode of assembly of the daughter discs and plane of the division is compatible with maintenance of the left-right asymmetry of the Giardia cytoskeleton in progeny, which cannot be satisfactorily explained by alternative models proposed so far.
Cell Motility and the Cytoskeleton 05/2007; 64(4):288-98. · 4.19 Impact Factor
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ABSTRACT: We studied morphological and molecular polymorphism of 53 Tetratrichomonas isolates obtained from amphibian, reptilian, mammalian hosts, and from a slug with the aid of protargol staining and analyses of ITS1-5.8S rRNA-ITS2, SSU rRNA, and alpha-tubulin gene sequences. The phylogenetic tree based on the concatenate of all sequences showed the monophyly of the genus Tetratrichomonas with respect to the genus Trichomonas. Our data suggest that two parabasalid genera, Pentatrichomonoides and Trichomonoides, may belong to the genus Tetratrichomonas. Tetratrichomonas isolates were divided into 16 robust host-specific and monophyletic groups that probably represent separate, mostly new, species. As only five Tetratrichomonas species were described from the examined host taxa so far, our study uncovered considerable species diversity within the genus. The wide host range, high level of species-specific host specificity, and newly revealed biodiversity make the genus Tetratrichomonas a valuable model for studying evolution of parasites.
Molecular Phylogenetics and Evolution 06/2006; 39(2):542-51. · 3.61 Impact Factor
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ABSTRACT: Giardia intestinalis is a binucleated diplomonad possessing four pairs of flagella of distinct location and function. Its pathogenic potential depends on the integrity of a complex microtubular cytoskeleton that undergoes a profound but poorly understood reorganization during cell division. We examined the cell division of G. intestinalis with the aid of light and electron microscopy and immunofluorescence methods and present here new observations on the reorganization of the flagellar apparatus in the dividing Giardia. Our results demonstrated the presence of a flagellar maturation process during which the flagella migrate, assume different position, and transform to different flagellar types in progeny until their maturation is completed. For each newly assembled flagellum it takes three cell cycles to become mature. The mature flagellum of Giardia is the caudal one that possesses a privileged basal body at which the microtubules of the adhesive disk nucleate. In contrast to generally accepted assumption that each of the two diplomonad mastigonts develops separately, we found that they are developmentally linked, exchanging their cytoskeletal components at the early phase of mitosis. The presence of the flagellar maturation process in a metamonad protist Giardia suggests that the basal body or centriole maturation is a universal phenomenon that may represent one of the core processes in a eukaryotic cell.
Eukaryotic Cell 05/2006; 5(4):753-61. · 3.60 Impact Factor
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ABSTRACT: Metronidazole and related 5-nitroimidazoles are the only available drugs in the treatment of human urogenital trichomoniasis caused by the protozoan parasite Trichomonas vaginalis. The drugs are activated to cytotoxic anion radicals by their reduction within the hydrogenosomes. It has been established that electrons required for metronidazole activation are released from pyruvate by the activity of pyruvate:ferredoxin oxidoreductase and transferred to the drug by a low-redox-potential carrier, ferredoxin. Here we describe a novel pathway involved in the drug activation within the hydrogenosome. The source of electrons is malate, another major hydrogenosomal substrate, which is oxidatively decarboxylated to pyruvate and CO2 by NAD-dependent malic enzyme. The electrons released during this reaction are transferred from NADH to ferredoxin by NADH dehydrogenase homologous to the catalytic module of mitochondrial complex I, which uses ferredoxin as electron acceptor. Trichomonads acquire high-level metronidazole resistance only after both pyruvate- and malate-dependent pathways of metronidazole activation are eliminated from the hydrogenosomes.
Antimicrobial Agents and Chemotherapy 01/2006; 49(12):5033-6. · 4.84 Impact Factor
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ABSTRACT: The protozoan genus Cochlosoma includes parasitic intestinal flagellates of birds and mammals of uncertain taxonomic classification. The presence of an adhesive disc, superficially similar to that of Giardia, led to a proposal that Cochlosoma should be classified as diplomonads. Careful morphological and ultrastructural observations, however, revealed conspicuous homologies to trichomonads. We addressed the question of classification and phylogenetic affiliation of Cochlosoma using the methods of molecular phylogenetics. Analyses based on the 16S rRNA gene sequence of the species Cochlosoma anatis very robustly placed Cochlosoma in the clade of the parabasalid subfamilies Trichomonadinae, Trichomitopsiinae and Pentatrichomonoidinae of the order Trichomonadida (bootstraps >94 %). The data did not provide robust support for any particular position of Cochlosoma within this clade because the sequence suffered from mutational saturation and produced a long branch. The most probable sister taxon of Cochlosoma is the genus Pentatrichomonas, because their relationship was supported specifically by the slowest-mutating, least-saturated positions as determined using the method slow-fast. Classification of the order Trichomonadida was revised to accommodate knowledge about its phylogeny - the family Cochlosomatidae and subfamilies Trichomitopsiinae and Pentatrichomonoidinae were abandoned, Trichomonadidae was amended and new families Tritrichomonadidae (formerly a subfamily) and Trichomitidae were proposed.
International journal of systematic and evolutionary microbiology 01/2006; 56(Pt 1):305-12. · 2.27 Impact Factor
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ABSTRACT: Circumscribing major eukaryote groups and resolving higher order relationships between them are among the most challenging tasks facing molecular evolutionists. Recently, evidence suggesting a new supergroup (the Excavata) comprising a wide array of flagellates has been collected. This group consists of diplomonads, retortamonads, Carpediemonas, heteroloboseans, Trimastix, jakobids, and Malawimonas, all of which possess a particular type of ventral feeding groove that is proposed to be homologous. Euglenozoans, parabasalids, and oxymonads have also been associated with Excavata as their relationships to one or more core excavate taxa were demonstrated. However, the main barrier to the general acceptance of Excavata is that its existence is founded primarily on cytoskeletal similarities, without consistent support from molecular phylogenetics. In gene trees, Excavata are typically not recovered together. In this paper, we present an analysis of the phylogenetic position of oxymonads (genus Monocercomonoides) based on concatenation of eight protein sequences (alpha-tubulin, beta-tubulin, gamma-tubulin, EF-1alpha, EF-2, cytosolic (cyt) HSP70, HSP90, and ubiquitin) and 18S rRNA. We demonstrate that the genes are in conflict regarding the position of oxymonads. Concatenation of alpha- and beta-tubulin placed oxymonads in the plant-chromist part of the tree, while the concatenation of other genes recovered a well-supported group of Metamonada (oxymonads, diplomonads, and parabasalids) that branched weakly with euglenozoans--connecting all four excavates included in the analyses and thus providing conditional support for the existence of Excavata.
Molecular Biology and Evolution 01/2006; 22(12):2508-18. · 5.55 Impact Factor
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ABSTRACT: Unikaryotic enteromonads and diplokaryotic diplomonads have been regarded as closely related protozoan groups. It has been proposed that diplomonads originated within enteromonads in a single event of karyomastigont duplication. This paper presents the first study to address these questions using molecular phylogenetics. The sequences of the small-subunit rRNA genes for three isolates of enteromonads were determined and a tree constructed with available diplomonad, retortamonad and Carpediemonas sequences. The diplomonad sequences formed two main groups, with the genus Giardia on one side and the genera Spironucleus, Hexamita and Trepomonas on the other. The three enteromonad sequences formed a clade robustly situated within the diplomonads, a position inconsistent with the original evolutionary proposal. The topology of the tree indicates either that the diplokaryotic cell of diplomonads arose several times independently, or that the monokaryotic cell of enteromonads originated by secondary reduction from the diplokaryotic state.
International journal of systematic and evolutionary microbiology 08/2005; 55(Pt 4):1729-33. · 2.27 Impact Factor
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ABSTRACT: Tetratrichomonas gallinarum is a widespread intestinal parasite of galliform and anseriform birds. The pathogenicity of this species is controversial, presenting an unsettled problem as yet. We analysed the polymorphism and genetic relationship among 29 isolates of T. gallinarum obtained from eight bird species and five T. gallinarum-like isolates from the oral cavity and lower respiratory tract of human patients. Two methods were used for the analyses: RAPD and sequencing of 16S rRNA, 5.8S rRNA, ITS1 and ITS2 genes, both producing consistent and well-supported results. The isolates were divided into five groups, A-E, with eleven subgroups. The distance between groups E, D and the cluster A-B-C considerably exceeded usual intraspecific polymorphism seen in trichomonads. Moreover, the largest subgroup, A2 (containing 18 isolates), was divided into three branches according to the host specificity. All isolates from humans were placed into avian subgroups A2 and B2. We conclude that our isolates represent, at least, three morphospecies or rather complexes of several cryptic species. Since certain species of the T. gallinarum complex can differ in their biological characteristics and some of them can infect humans, the problem of T. gallinarum pathogenicity should be re-examined with regard to specific genetic groups and zoonotic potential of some of these lineages should be considered.
Veterinary Parasitology 04/2005; 128(1-2):11-21. · 2.58 Impact Factor
