Adhesion of Tritrichomonas foetus to bovine vaginal epithelial cells.
An in vitro culture system of bovine vaginal epithelial cells (BVECs) was developed to study the cytopathogenic effects of Tritrichomonas foetus and the role of lipophosphoglycan (LPG)-like cell surface glycoconjugates in adhesion of parasites to host cells. Exposure of BVEC monolayers to T. foetus resulted in extensive damage of monolayers. Host cell disruption was measured quantitatively by a trypan blue exclusion assay and by release of (3)H from [(3)H]thymidine-labeled host cells. Results indicated contact-dependent cytotoxicity of host cells by T. foetus. The cytopathogenic effect was a function of T. foetus density. Metronidazole- or periodate-treated T. foetus showed no damage to BVEC monolayers. A related human trichomonad, Trichomonas vaginalis, showed no cytotoxic effects, indicating species-specific host-parasite interactions. A direct binding assay was developed and used to investigate the role of a major cell surface LPG-like molecule in host-parasite adhesion. The results of competition experiments showed that the binding to BVECs was displaceable, was saturable, and yielded a typical binding curve, suggesting that specific receptor-ligand interactions mediate the attachment of T. foetus to BVECs. Progesterone-treated BVECs showed enhanced parasite binding. T. foetus LPG inhibited the binding of T. foetus to BVECs; the LPG from T. vaginalis and a variety of other glycoconjugates did not. These data imply specificity of LPG on host-parasite adhesion. Periodate-treated parasites showed no adherence to host cells, indicating the involvement of carbohydrate containing molecules in the adhesion process. (+info)
Cytopathogenic effect of Trichomonas vaginalis on human vaginal epithelial cells cultured in vitro.
In this study we established human vaginal epithelial cells (hVECs) in culture and evaluated their interaction with Trichomonas vaginalis parasites to complement previous studies using other cell types. Primary cultures of hVECs were established. Contaminating fibroblasts were separated from epithelial cells by differential trypsinization. Specific antibody staining revealed that over 92% of cells in hVEC monolayers were epithelial cells. T. vaginalis adhered to hVECs and produced severe cytotoxic effects resulting in obliteration of the monolayer within 24 h. Adherence and cytotoxicity were not observed when T. vaginalis was exposed to human vaginal fibroblasts or bovine vaginal epithelial cells. Likewise, the bovine parasite Tritrichomonas foetus had no cytotoxic effects on hVECs. We concluded that the interaction between T. vaginalis and hVECs is both cell specific (limited to epithelial cells and not vaginal fibroblasts) and species specific (limited to human vaginal cells and not bovine cells). Pretreatment of T. vaginalis with metronidazole or periodate abolished the adhesion of parasites to cell monolayers and the cytotoxic effect, suggesting involvement of carbohydrate-containing molecules in these processes. Different clinical isolates of T. vaginalis caused damage to cultured cells at different rates. Parasites separated from the vaginal cell monolayer by a permeable membrane did not produce a cytopathic effect, suggesting contact-dependent cytotoxicity. (+info)
Strategies by which some pathogenic trichomonads integrate diverse signals in the decision-making process.
The interaction between each one of Trichomonas vaginalis and Tritrichomonas foetus with their hosts is a complex process in which components associated to the cell surfaces of both parasites and host epithelial cells, and also to soluble components found in vaginal/urethral secretions, are involved. Either cytoadhesion or the cytotoxicity exerted by parasites to host cells can be dictated by virulence factors such as adhesins, cysteine proteinases, laminin-binding proteins, integrins, integrin-like molecules, a cell detachment factor, a pore-forming protein, and glycosidases among others. How trichomonads manipulate informations from the extracellular medium, transduce such informations, and respond to them by stimulating the activities of some surface molecules and/or releasing enzymes are the aspects concerning trichomonal virulence which are here briefly reviewed and discussed. (+info)
Morphologic aspects of Tetratrichomonas didelphidis isolated from opossums Didelphis marsupialis and Lutreolina crassicaudata.
Tetratrichomonas didelphidis (Hegner & Ratcliffe, 1927) Andersen & Reilly, 1965 is a flagellate protozoan found in the intestine, cecum, and colon of Didelphis marsupialis. The parasitic protozoa used in this study was found and isolated in the intestine of opossums in Pavlova starch-containing medium in Florianopolis, State of Santa Catarina, Brazil, from D. marsupialis and Lutreolina crassicaudata. The strains were cultivated in Diamond medium without maltose and with starch solution, pH 7.5 at 28 degrees C. The specimens were stained by the Giemsa method and Heidenhain's iron hematoxylin. The light microscopy study of the trophozoites revealed the same morphologic characteristics as specimens previously described. (+info)
Construction and bootstrap analysis of DNA fingerprinting-based phylogenetic trees with the freeware program FreeTree: application to trichomonad parasites.
The Win95/98/NT program FreeTree for computation of distance matrices and construction of phylogenetic or phenetic trees on the basis of random amplified polymorphic DNA (RAPD), RFLP and allozyme data is presented. In contrast to other similar software, the program FreeTree (available at http://www.natur.cuni.cz/~flegr/programs/freetree or http://ijs.sgmjournals.org/content/vol51/issue3/) can also assess the robustness of the tree topology by bootstrap, jackknife or operational taxonomic unit-jackknife analysis. Moreover, the program can be also used for the analysis of data obtained in several independent experiments performed with non-identical subsets of taxa. The function of the program was demonstrated by an analysis of RAPD data from 42 strains of 10 species of trichomonads. On the phylogenetic tree constructed using FreeTree, the high bootstrap values and short terminal branches for the Tritrichomonas foetus/suis 14-strain branch suggested relatively recent and probably clonal radiation of this species. At the same time, the relatively lower bootstrap values and long terminal branches for the Trichomonas vaginalis 20-strain branch suggested more ancient radiation of this species and the possible existence of genetic recombination (sexual reproduction) in this human pathogen. The low bootstrap values and the star-like topology of the whole Trichomonadidae tree confirm that the RAPD method is not suitable for phylogenetic analysis of protozoa at the level of higher taxa. It is proposed that the repeated bootstrap analysis should be an obligatory part of any RAPD study. It makes it possible to assess the reliability of the tree obtained and to adjust the amount of collected data (the number of random primers) to the amount of phylogenetic signals in the RAPD data of the taxon analysed. The FreeTree program makes such analysis possible. (+info)
The classic approach to diagnosis of vulvovaginitis: a critical analysis.
OBJECTIVE: To correlate the symptoms, signs and clinical diagnosis in women with vaginal discharge, based on the combined weight of the character of the vaginal discharge and bedside tests, with the laboratory diagnosis. METHODS: Women presenting consecutively to the women's health center with vaginal discharge were interviewed and examined for assessment of the quantity and color of the discharge. One drop of the material was then examined for pH and the whiff test was done; a wet mount in saline and in 10% KOH was examined microscopically. The clinical diagnosis was based on the results of these assessments. Gram stain and cultures of the discharge were sent to the microbiology laboratory. RESULTS: One hundred and fifty-three women with vaginal discharge with a clinical diagnosis of vulvovaginitis participated in the study. Fifty-five (35.9%) had normal flora and the other 98 (64.1%) had true infectious vulvovaginitis (kappa agreement = 18%). According to the laboratory, the principal infectious micro-organism causing the vulvovaginitis was Candida species. Candida infection was associated with pH levels of less than 4.5 (p < 0.0001, odds ratio = 4.74, 95% confidence interval: 2.35-9.5, positive predictive value 68.4%). The whiff test was positive in only a small percentage of bacterial vaginosis (BV) (p = not significant (NS)). Clue cells were documented in 53.3% of patients with a laboratory diagnosis of BV (p < 0.02, positive predictive value 26.7%). CONCLUSIONS: The current approach to the diagnosis of vulvovaginitis should be further studied. The classical and time-consuming assessments were shown not to be reliable diagnostic measures. (+info)
Reconstructing/deconstructing the earliest eukaryotes: how comparative genomics can help.
We could reconstruct the evolution of eukaryote-specific molecular and cellular machinery if some living eukaryotes retained primitive cellular structures and we knew which eukaryotes these were. It's not clear that either is the case, but the expanding protist genomic database could help us in several ways. (+info)
Application of a PCR assay to enhance the detection and identification of Tritrichomonas foetus in cultured preputial samples.
The traditional diagnostic test for Tritrichomonas foetus involves collection of preputial or vaginal samples followed by culture in a growth media and microscopic examination. Recently, polymerase chain reaction (PCR) techniques have been described for use as a diagnostic assay. The objective of this study was to evaluate a previously described PCR assay for detecting T. foetus in cultured preputial material. The detection limits of the assay for T. foetus organisms in a growth medium, in samples prepared from washing microscope slides, and in preputial material cultured in a growth medium were determined. Preputial samples were collected from 13 bulls uninfected with T. foetus. The PCR assay was able to detect 5 T. foetus organisms in the growth medium and the cultured preputial material. Amplification products were obtained from samples prepared from washes of microscope slides containing as few as 3 visualized organisms. The PCR assay was able to detect organisms in culture at a lower concentration than was possible by direct microscopic examination. This low detection limit may allow the PCR assay to be used to enhance the sensitivity of the current diagnostic test. In addition, the assay could be used to confirm the identification of T. foetus organisms observed by direct microscopic examination when other confirmation techniques, such as staining and phase microscopy, are not practical. (+info)