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PubMed1988

Multiple populations of double-stranded RNA in two virus-harbouring strains of Trichomonas vaginalis.

The existence of six dsRNA segments of Trichomonas vaginalis virus was confirmed and the molar mass and relative abundance of these segments were determined by agarose gel electrophoresis with reovirus dsRNA serving as a standard. The M's were 3.5, 3.4, 3.2, 2.5, 1.4 and 0.34 Mg/mol for the two strains studied, the relative abundances, however, were 1.0, 1.4, 3.0, 0.3, 2.7, 4.2 and 1.0, 0.6, 1.7, 0.5, 3.4 1.0 for these strains, respectively. Cell homogenate fractionation showed that all dsRNA segments were associated with viral particles. The data appeared to support the hypothesis of a relationship between viruses of the protozoan T. vaginalis and of the yeast Saccharomyces cerevisiae.

Animals↗
PubMed1997

Phylogenetic position of the mitochondrion-lacking protozoan Trichomonas tenax, based on amino acid sequences of elongation factors 1alpha and 2.

Major parts of amino-acid-coding regions of elongation factor (EF)-1alpha and EF-2 in Trichomonas tenax were amplified by PCR from total genomic DNA and the products were cloned into a plasmid vector, pGEM-T. The three clones from each of the products of the EF-1alpha and EF-2 were isolated and sequenced. The insert DNAs of the clones containing EF-1alpha coding regions were each 1,185 bp long with the same nucleotide sequence and contained 53.1% of G + C nucleotides. Those of the clones containing EF-2 coding regions had two different sequences; one was 2,283 bp long and the other was 2,286 bp long, and their G + C contents were 52.5 and 52.9%, respectively. The copy numbers of the EF-1alpha and EF-2 gene per chromosome were estimated as four and two, respectively. The deduced amino acid sequences obtained by the conceptual translation were 395 residues from EF-1alpha and 761 and 762 residues from the EF-2s. The sequences were aligned with the other eukaryotic and archaebacterial EF-1alphas and EF-2s, respectively. The phylogenetic position of T. tenax was inferred by the maximum likelihood (ML) method using the EF-1alpha and EF-2 data sets. The EF-1alpha analysis suggested that three mitochondrion-lacking protozoa, Glugea plecoglossi, Giardia lamblia, and T. tenax, respectively, diverge in this order in the very early phase of eukaryotic evolution. The EF-2 analysis also supported the divergence of T. tenax to be immediately next to G. lamblia.

Amino Acid Sequence↗
PubMed1998

The pyrophosphate-dependent phosphofructokinase of the protist, Trichomonas vaginalis, and the evolutionary relationships of protist phosphofructokinases.

The pyrophosphate-dependent phosphofructokinase (PPi-PFK) of the amitochondriate protist Trichomonas vaginalis has been purified. The enzyme is a homotetramer of about 50 kDa subunits and is not subject to allosteric regulation. The protein was fragmented and a number of peptides were sequenced. Based on this information a PCR product was obtained from T. vaginalis gDNA and used to isolate corresponding cDNA and gDNA clones. Southern analysis indicated the presence of five genes. One open reading frame (ORF) was completely sequenced and for two others the 5' half of the gene was determined. The sequences were highly similar. The complete ORF corresponded to a polypeptide of about 46 kDa. All the peptide sequences obtained were present in the derived sequences. The complete ORF was highly similar to that of other PFKs, primarily in its amino-terminal half. The T. vaginalis enzyme was most similar to PPi-PFK of the mitochondriate heterolobosean, Naegleria fowleri. Most of the residues shown or assumed to be involved in substrate binding in other PPi-PFKs were conserved in the T. vaginalis enzyme. Direct comparison and phylogenetic reconstruction revealed a significant divergence among PPi-PFKs and related enzymes, which can be assigned to at least four distantly related groups, three of which contain enzymes of protists. The separation of these groups is supported with a high percentage of bootstrap proportions. The short T. vaginalis PFK shares a most recent common ancestor with the enzyme from N. fowleri. This pair is clearly separated from a group comprising the long (>60-kDa) enzymes from Giardia lamblia, Entamoeba histolytica pfk2, the spirochaetes Borrelia burgdorferi and Trepomena pallidum, as well as the alpha- and beta-subunits of plant PPi-PFKs. The third group ("X") containing protist sequences includes the glycosomal ATP-PFK of Trypanosoma brucei, E. histolytica pfk1, and a second sequence from B. burgdorferi. The fourth group ("Y") comprises cyanobacterial and high-G + C, Gram-positive eubacterial sequences. The well-studied PPi-PFK of Propionibacterium freudenreichii is highly divergent and cannot be assigned to any of these groups. These four groups are well separated from typical ATP-PFKs, the phylogenetic analysis of which confirmed relationships established earlier. These findings indicate a complex history of a key step of glycolysis in protists with several early gene duplications and possible horizontal gene transfers.

Amino Acid Sequence↗
PubMed2000

Immunolocalization of two hydrogenosomal enzymes of Trichomonas vaginalis.

Three monoclonal antibodies specific for malic enzyme and for the alpha- and beta-subunits, respectively, of the succinyl-coenzyme A (CoA) synthetase of Trichomonas vaginalis were used to immunolocalize these proteins in the cell. All antibodies labeled the hydrogenosome matrix as determined both by immunofluorescence and by immunogold staining. There was no labeling on the cell surface or in any other cell compartment. These results support the idea that these proteins are restricted to a hydrogenosomal function and do not play a role as adhesins at the plasma membrane surface.

Animals↗
PubMed2004

Trichomonas vaginalis degrades nitric oxide and expresses a flavorubredoxin-like protein: a new pathogenic mechanism?

Besides possessing many physiological roles, nitric oxide (NO) produced by the immune system in infectious diseases has antimicrobial effects. Trichomoniasis, the most widespread non-viral sexually transmitted disease caused by the microaerophilic protist Trichomonas vaginalis, often evolves into a chronic infection, with the parasite able to survive in the microaerobic, NO-enriched vaginal environment. We relate this property to the finding that T. vaginalis degrades NO under anaerobic conditions, as assessed amperometrically. This activity, which is maximal (133 +/- 41 nmol NO/10(8) cells per minute at 20 degrees C) at low NO concentrations (< or = 1.2 microM), was found to be: (i) NADH dependent, (ii) cyanide insensitive and (iii) inhibited by O(2). These features are consistent with those of the Escherichia coli A-type flavoprotein (ATF), recently discovered to be endowed with NO reductase activity. Using antibodies against the ATF from E. coli, a protein band was immunodetected in the parasite grown in a standard medium. If confirmed, the expression of an ATF in eukaryotes suggests that the genes coding for ATFs were transferred during evolution from anaerobic Prokarya to pathogenic protists, to increase their fitness for the microaerobic, parasitic life style. Thus the demonstration of an ATF in T. vaginalis would appear relevant to both pathology and evolutionary biology. Interestingly, genomic analysis has recently demonstrated that Giardia intestinalis and other pathogenic protists have genes coding for ATFs.

Animals↗
PubMed2001

Mycoplasma hominis and Trichomonas vaginalis symbiosis: multiplicity of infection and transmissibility of M. hominis to human cells.

We recently reported that most Trichomonas vaginalis isolates cultured in vitro are infected by Mycoplasma hominis. In this work, we have characterized some aspects of the relationships between the two microorganisms. PCR, cultivation, and immunological methods revealed that the number of M. hominis organisms carried by T. vaginalis in culture varied from isolate to isolate, suggesting a specific multiplicity of infection. Moreover, infected T. vaginalis isolates were able to pass bacteria not only to M. hominis-free protozoa, but also to human-derived epithelial cells. The in vitro transmission of the bacterium from T. vaginalis to both uninfected parasite isolates and human epithelial cells suggests a role for T. vaginalis as a carrier of the M. hominis infection in vivo.

Animals↗
PubMed2004

Comparative analysis of the ribosomal components of the hydrogenosome-containing protist, Trichomonas vaginalis.

The ribosomes of the amitochondriate but hydrogenosome-containing protist lineage, the trichomonads, have previously been reported to be prokaryotic or primitive eukaryotic, based on evidence that they have a 70S sedimentation coefficient and a small number of proteins, similar to prokaryotic ribosomes. In order to determine whether the components of the trichomonad ribosome indeed differ from those of typical eukaryotic ribosomes, the ribosome of a representative trichomonad, Trichomonas vaginalis, was characterized. The sedimentation coefficient of the T. vaginalis ribosome was smaller than that of Saccharomyces cerevisiae and larger than that of Escherichia coli. Based on two-dimensional PAGE analysis, the number of different ribosomal proteins was estimated to be approximately 80. This number is the same as those obtained for typical eukaryotes (approximately 80) but larger than that of E. coli (approximately 55). N-Terminal amino acid sequencing of 18 protein spots and the complete sequences of 4 ribosomal proteins as deduced from their genes revealed these sequences to display typical eukaryotic features. Phylogenetic analyses of the five ribosomal proteins currently available also clearly confirmed that the T. vaginalis sequences are positioned within a eukaryotic clade. Comparison of deduced secondary structure models of the small and large subunit rRNAs of T. vaginalis with those of other eukaryotes revealed that all helices commonly found in typical eukaryotes are present and conserved in T. vaginalis, while variable regions are shortened or lost. These lines of evidence demonstrate that the T. vaginalis ribosome has no prokaryotic or primitive eukaryotic features but is clearly a typical eukaryotic type.

Amino Acid Sequence↗
PubMed2002

Scanning electron microscopy in the investigation of the in vitro hemolytic activity of Trichomonas vaginalis.

The in vitro hemolytic activity of Trichomonas vaginalis has been previously demonstrated, but the mechanisms involved remain to be elucidated. In this work we used scanning electron microscopy to investigate the contact dependency of the hemolytic phenomenon caused by the parasites. The erythrocytes adhered to the parasites' surface and were phagocytosed. These observations suggest that the contact between T. vaginalis and erythrocytes may be an important mechanism in the injury caused to the erythrocytes. The hemolytic activity of T. vaginalis may be an efficient means of obtaining nutrients for the parasite and allow the investigation of the mechanism used by T. vaginalis to damage cellular membranes.

Animals↗
PubMed2002

Characterization of an ecto-phosphatase activity in the human parasite Trichomonas vaginalis.

We have characterized phosphatase activity present on the external surface of Trichomonas vaginalis, using intact living parasites. This enzyme hydrolyzes the substrate p-nitrophenylphosphate (p-NPP) at a rate of 134.3+/-14.8 nmol Pi/h per 10(7) cells. This phosphatase activity decreased by increasing the pH from 6.8 to 8.4, a pH range in which cell viability was maintained for at least 1 h. Experiments using classical inhibitors of acid phosphatases, such as ammonium molybdate and sodium fluoride, as well as inhibitors of phosphotyrosine phosphatase, such as sodium orthovanadate, [monoperoxo(picolinato)oxovanadate(V)] (mpV-PIC) and [potassiumbisperoxo(1,10-phenanthroline)oxovanadate(V)] (bpV-PHEN), showed a decrease in this phosphatase activity, with different patterns of inhibition. Cytochemical analysis showed the localization of this enzyme on the parasite surface (cell body and flagellum) and in intracellular vacuoles. Phosphatase reaction products were also observed in exocytosed membrane-bound material.

4-Nitrophenylphosphatase↗
PubMed2003

Morphogenesis during division and griseofulvin-induced changes of the microtubular cytoskeleton in the parasitic protist, Trichomonas vaginalis.

The behavior of microtubular structures during division was followed by immunofluorescence in Trichomonas vaginalis using an anti-alpha-tubulin monoclonal antibody together with nuclear staining by DAPI, allowing us to describe successive mitotic stages. In contrast to recent reports, we showed that: (1) the microtubular axostyle-pelta complex depolymerized during division, (2) the flagella were assembled during mitosis, and (3) the flagellar number was restored in each daughter kinetid before cytokinesis. Observation of griseofulvin-treated T. vaginalis cells revealed that the elongation of the mitotic spindle or paradesmosis was not the main motile force separating the daughter kinetids to opposite poles during division, suggesting the existence of other mechanisms and/or molecules involved in this morphogenetic event. Examination of treated cells re-incubated in fresh medium showed the nucleation of microtubules radiating from the perinuclear area, the origin of which is discussed. Finally, we confirm the effectiveness of griseofulvin against T. vaginalis and propose that this antifungal drug could be a promising antitrichomonal agent.

Animals↗
PubMed2004

Trichomonas vaginalis: cytochemical localization of a NTPDase1 and an ecto-5'-nucleotidase and effects of adenine nucleotides on cellular viability.

Nucleoside triphosphate diphosphohydrolase 1 (NTPDase1), which hydrolyzes extracellular ATP and ADP, and ecto-5'-nucleotidase, which hydrolyzes AMP, are characterized for Trichomonas vaginalis. Ultrastructural cytochemical microscopy showed NTPDase1 and ecto-5'-nucleotidase activities on the surface of the parasites. High levels of extracellular adenine nucleotides and adenosine did not exert cytolytic effects in intact cells of T. vaginalis. Our results suggest that these enzymes are relevant for the survival of the parasite during exposure to extracellular nucleotides. Since the ecto-localization of these enzymes is essential for the maintenance of adenosine extracellular levels, this nucleoside could be important for the purine salvage pathway in the parasite.

5'-Nucleotidase↗
PubMed2004

Trichomonas vaginalis virulence against epithelial cells and morphological variability: the comparison between a well-established strain and a fresh isolate.

The FMVI strain of Trichomonas vaginalis was freshly isolated from an asymptomatic patient, and its morphological properties and virulence in vitro compared with the well-established JT strain. The morphological variability of the parasites was assessed by differential interference microscopy and both scanning and transmission electron microscopy. The FMV1 strain presented nearly 20% amoeboid cells whereas the JT strain presented high percentages of ellipsoid but no amoeboid cells. The FMV1 morphotype population was unaltered after at least 1 year of subculturing. Electron microscopy revealed that this strain produced numerous pseudopod structures which mediated intimate contact and interdigitation among trophozoites. Dead FMV1 parasites were often phagocytosed by conspecific cells. We also compared the cytolytic capacity of these two populations against epithelial MDCK cells and its contact dependence. The FMV1 strain rapidly adhered to plastic or glass surfaces and to MDCK monolayers. This strain destroyed about 93% of the epithelial cells in 90 min whereas the cytolytic activity of the JT parasites was very much lower (about 41%). Parasite supernatants displayed no cytolytic activity, indicating contact-mediated lysis. The protozoan virulence in vitro did not correlate well with the clinical observations. The implications of these results are discussed.

Animals↗
PubMed2006

Symbiosis of Mycoplasma hominis in Trichomonas vaginalis may link metronidazole resistance in vitro.

Fourteen of 28 Trichomonas vaginalis isolates collected from patients in Guangzhou, China from 2003 to 2004 were found to be naturally infected with Mycoplasma hominis, as determined by PCR using specific primers. In vitro metronidazole sensitivity assay of the 28 isolates revealed four displaying low susceptibility [minimum lethal concentration (MLC)= approximately 13-25 microg/ml] and another four displaying high resistance (MLC=50-100 microg/ml). The overwhelming majority of these resistant isolates (7/8) were mycoplasma-infected. The mean of MLCs of mycoplasma-infected isolates is approximately 10-fold higher than the mean of noninfected isolates (p=0.029). Sequence analyses of PCR-amplified small subunit-large subunit rRNA interspacer regions (ITS1/5.8S/ITS2) revealed that 23 of the 28 samples are identical, the remaining five being separable into two groups, each with a single point mutation. These internal transcribed spacer sequence variants are associated neither with mycoplasma infection nor with drug resistance. In contrast, random amplified polymorphic DNA analyses of DNAs using 10 different primers showed that the drug-resistant isolates are clustered together in association with mycoplasma infection, albeit more loosely. Taken together, the results obtained from this study suggest that in vitro metronidazole resistance of T. vaginalis is related to mycoplasma infection of this protozoan.

Animals↗
PubMed2006

Antiparasitic activity of two Lavandula essential oils against Giardia duodenalis, Trichomonas vaginalis and Hexamita inflata.

Two essential oils derived from Lavandula angustifolia and Lavandula x intermedia were investigated for any antiparasitic activity against the human protozoal pathogens Giardia duodenalis and Trichomonas vaginalis and the fish pathogen Hexamita inflata: all of which have significant infection and economic impacts. The study has demonstrated that low (< or = 1%) concentrations of L. angustifolia and L. x intermedia oil can completely eliminate T. vaginalis, G. duodenalis and H. inflata in vitro. At 0.1% concentration, L. angustifolia oil was found to be slightly more effective than L x intermedia oil against G. duodenalis and H. inflata. The potential applications are discussed.

Animals↗
PubMed2001

Hydrogenosome morphological variation induced by fibronectin and other drugs in Trichomonas vaginalis and Tritrichomonas foetus.

The hydrogenosome is a spherical organelle, found in some anaerobic protozoa, which participates in ATP and molecular hydrogen formation. The morphological alterations in hydrogenosomes induced by fibronectin, hydroxyurea and cytochalasin B in Trichomonas vaginalis and Tritrichomonas foetus are presented. We demonstrate that, under experimental conditions, the hydrogenosome presents a high diversity in size and shape, suggesting a mechanism that seems to compensate for the stress provoked by drugs. The following experimental procedures were used: (1) fibronectin-mediated endocytic activity, (2) 4 mM hydroxyurea for 15 h, and (3) 10 microg cytochalasin B/ml in the culture medium. The main alterations observed in hydrogenosomes were: (1) formation of giant hydrogenosomes, (2) presence of internal membranes, (3) increased diversity of non-spherical forms, some of them bizarre, (4) presence of sub-compartments in the matrix, as vesicles, (5) presence of ribosome-like particles on the outer hydrogenosomal membrane, (6) enlargement of the peripheral vesicle, and (7) continuity with membrane profiles.

Animals↗
PubMed1998

Observation of membrane fusion on the interaction of Trichomonas vaginalis with human vaginal epithelial cells.

The in vitro cytopathic effect of Trichomonas vaginalis on epithelial cells was analyzed through the interaction of two parasite strains with freshly collected human vaginal epithelial cells (HVECs) from normal women. Videomicroscopy, light and electron microscopy (scanning and transmission), freeze-fracture, the tracer lanthanum nitrate, and the periodic acid-thio-semicarbazide-silver proteinate techniques were used to analyze regions of close contact between the HVECs and T. vaginalis. After 2 h of interaction, all HVECs were dead, whereas all the trichomonads were alive. Microscopic observations demonstrated that in addition to previously described regions of adhesion and interdigitations, areas of continuity between the cytoplasm of the two interacting cells were found. They were not easy to find since they correspond to focal spots placed in different depths of the section. When these regions were depicted, the plasma membranes of the T. vaginalis and the vaginal epithelial cells seemed to be fused.

Animals↗
PubMed1998

Involvement of Trichomonas vaginalis surface-associated glycoconjugates in the parasite/target cell interaction. A quantitative electron microscopy study.

Cytochemical labeling with gold particle-conjugated lectins in combination with transmission and scanning electron microscopy was used to localise specific sugar residues on the Trichomonas vaginalis cell surface. For investigation of the role played by the surface glycoconjugates of T. vaginalis in the process of parasite adhesion to the target cells, selected glycan moieties of parasite surface-bound molecules were removed by treatment with alpha-mannosidase and beta-N-acetylglucosaminidase. For observation of the parasite/epithelial cell interaction, human amnion membrane was employed as an in vitro model. Ultrastructure observations showed that T. vaginalis has distinct binding sites for concanavalin-A and wheat-germ agglutinin. This indicates the presence of mannose or mannose-like residues and N-acetyl-D-glucosamine-containing residues on the parasite membrane. The addition of inhibitory sugars to T. vaginalis incubation media diminished the subsequent labeling of the parasite cell coat with lectins. Enzyme treatment caused a significant reduction in the number of sugar residues on the cell surface of the parasite. The majority of the viable, motile, enzyme-treated T. vaginalis cells incubated with amnion membrane were incapable of adhering to the target cells. It was concluded that sugar residues, in particular alpha-D-mannose and N-acetyl-glucosamine, present in the parasite glycocalyx are involved in the process of T. vaginalis attachment to the host's epithelial cells. Removal of the T. vaginalis cell-surface sugars prevented the attachment to and damage of the epithelial cells.

Acetylglucosaminidase↗
PubMed1998

Isolation and characterization of flagella from Trichomonas vaginalis.

As a first step in the biochemical and biomedical analyses of flagella from Trichomonas vaginalis the flagella were isolated, purified, and analyzed. The flagella were detached by mechanical shearing and a crude flagellar preparation was obtained by low-speed differential centrifugation. The crude flagellar preparation was subjected to further purification by discontinuous sucrose density-gradient centrifugation. Electron micrographs (EM) of the purified flagella showed the typical 9 + 2 axonemal arrangement. The structural integrity and the flagellar membrane were not destroyed by the deflagellation method or the purification scheme employed. The flagellar preparations were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The purified preparation contained many flagellar enriched proteins ranging from 20 to 120 kDa. Three major proteins of 65 kDa and a doublet of about 50-58 kDa were observed. The protein patterns and EM appearance of the fractions were highly reproducible.

Animals↗