CP30, a cysteine proteinase involved in Trichomonas vaginalis cytoadherence.
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We describe here the participation of a Trichomonas vaginalis 30-kDa proteinase (CP30) with affinity to the HeLa cell surface in attachment of this parasite to host epithelial cells. The CP30 band is a cysteine proteinase because its activity was inhibited by E-64, a thiol proteinase inhibitor. In two-dimensional substrate gel electrophoresis of total extracts of the trichomonad isolate CNCD 147, three spots with proteolytic activity were detected in the 30-kDa region, in the pI range from 4.5 to 5.5. Two of the spots (pI 4.5 and 5.0) bound to the surfaces of fixed HeLa cells corresponding to the CP30 band. The immunoglobulin G fraction of the rabbit anti-CP30 antiserum that recognized a 30-kDa band by Western blotting and immunoprecipitated CP30 specifically inhibited trichomonal cytoadherence to HeLa cell monolayers in a concentration-dependent manner and reacted with CP30 at the parasite surface. CP30 degraded proteins found on the female urogenital tract, including fibronectin, collagen IV, and hemoglobin. Interestingly, CP30 digested fibronectin and collagen IV only at pH levels between 4.5 and 5.0. Moreover, trichomonosis patients whose diagnosis was confirmed by in vitro culture possessed antibody to CP30 in both sera and vaginal washes, and CP30 activity was found in vaginal washes. Our results suggest that surface CP30 is a cysteine proteinase necessary for trichomonal adherence to human epithelial cells. (+info)
Non-specific urethritis and the tetracyclines.
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The possible aetiological factors in non-gonococcal and non-specific urethritis are reviewed. The response of NSU to various courses of different tetracycline drugs is assessed. Prolonged courses of treatment did not give better results than shorter courses. When reviewing the infective aetiology of non-gonococcal urethritis, it was noted that more than one organism (or potential pathogen) would be present in many cases. It is therefore surmised that there may be at times a mixed aetiology and at other times a truly non-specific aetiology. Isolations by different workers have indicated that the following organisms might be expected: Chlamydia 40 per cent.; Mycoplasma-M. hominis 20 per cent., T-strain over 60 per cent.; Trichomonas 15 per cent.; Candida possibly over 5 per cent. Truly non-specific urethritis may account for 25 to 30 per cent. of cases. (+info)
Microbial flora of the lower genital tract during pregnancy: relationship to morbidity.
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Nineteen genera and groups of micro-organisms were isolated from the lower genital tract of 280 women at their first antenatal visit. Chlamydia, viruses, and T-strain mycoplasmas were not sought, and only routine methods of anaerobic culture were used. Growth was recorded as scanty, moderate or heavy. The population studied was grouped according to age, parity, gestational stage at booking, presence and degree of severity of lower genital tract morbidity, past history of vulvovaginitis, and suspicion of lower genital tract morbidity as evidenced by a request for a report on the microbiological findings. The frequency of isolation of the various microbes in health and in disease is given. The grading of Gram-stained smears bore no relation to the isolation rates of lactobacilli, but there was a significant increase (p less than 0-001) in the isolation rates of each of the following: Mycoplasma hominis, Bacteroides spp., Trichomonas vaginalis, Gram-variable cocco-bacilli, and anaerobic streptococci in those patients with smears in which lactobacilli were adjudged to be absent. The isolation of faecal streptococci was increased (p less than 0-001) in women aged more than 34 years. Escherichia coli (p less than 0-05) and anaerobic and microaerophilic streptococci (p less than 0-02) were isolated more frequently from those booking after the 25th week of pregnancy. The incidence of M. hominis (p less than 0-02) and of anaerobic streptococci (p less than 0-05) increased between the first and third trimesters. No significance positive correlations were established between the isolation rates of the various microbes and objective assessment of lower genital tract morbidity or the demonstration of pus cells, but lactobacilli were isolated less frequently (p less than 0-01) from those with morbidity. The isolation of Candida albicans (p less than 0-02), T. vaginalis (p less than 0-05), and M. hominis (p less than 0.05) was increased in patients in whom vulvovaginitis was suspected, and that of T. vaginalis (p less than 0-05) was increased in those with a past history of vulvovaginitis. The study indicates that, other than the pathogens T. vaginalis and C. albicans, only M. Hominis could be suspected, on statistical grounds, of being associated with disease of the lower genital tract during early pregnancy. (+info)
Detection of trichomonosis in vaginal and urine specimens from women by culture and PCR.
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Vaginal trichomonosis is a highly prevalent infection which has been associated with human immunodeficiency virus acquisition and preterm birth. Culture is the current "gold standard" for diagnosis. As urine-based testing using DNA amplification techniques becomes more widely used for other sexually transmitted diseases (STDs) such as gonorrhea and chlamydia, a similar technique for trichomonosis would be highly desirable. Women attending an STD clinic for a new complaint were screened for Trichomonas vaginalis by wet-preparation (wet-prep) microscopy and culture and for the presence of T. vaginalis DNA by specific PCR of vaginal and urine specimens. The presence of trichomonosis was defined as the detection of T. vaginalis by direct microscopy and/or culture from either vaginal samples or urine. The overall prevalence of trichomonosis in the population was 28% (53 of 190). The sensitivity and specificity of PCR using vaginal samples were 89 and 97%, respectively. Seventy-four percent (38 of 51) of women who had a vaginal wet prep or vaginal culture positive for trichomonads had microscopic and/or culture evidence of the organisms in the urine. Two women were positive for trichomonads by wet prep or culture only in the urine. The sensitivity and specificity of PCR using urine specimens were 64 and 100%, respectively. These results indicate that the exclusive use of urine-based detection of T. vaginalis is not appropriate in women. PCR-based detection of T. vaginalis using vaginal specimens may provide an alternative to culture. (+info)
Origin and evolution of eukaryotic chaperonins: phylogenetic evidence for ancient duplications in CCT genes.
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Chaperonins are oligomeric protein-folding complexes which are divided into two distantly related structural classes. Group I chaperonins (called GroEL/cpn60/hsp60) are found in bacteria and eukaryotic organelles, while group II chaperonins are present in archaea and the cytoplasm of eukaryotes (called CCT/TriC). While archaea possess one to three chaperonin subunit-encoding genes, eight distinct CCT gene families (paralogs) have been characterized in eukaryotes. We are interested in determining when during eukaryotic evolution the multiple gene duplications producing the CCT subunits occurred. We describe the sequence and phylogenetic analysis of five CCT genes from TRICHOMONAS: vaginalis and seven from GIARDIA: lamblia, representatives of amitochondriate protist lineages thought to have diverged early from other eukaryotes. Our data show that the gene duplications producing the eight CCT paralogs took place prior to the organismal divergence of TRICHOMONAS: and GIARDIA: from other eukaryotes. Thus, these divergent protists likely possess completely hetero-oligomeric CCT complexes like those in yeast and mammalian cells. No close phylogenetic relationship between the archaeal chaperonins and specific CCT subunits was observed, suggesting that none of the CCT gene duplications predate the divergence of archaea and eukaryotes. The duplications producing the CCTdelta and CCTepsilon subunits, as well as CCTalpha, CCTbeta, and CCTeta, are the most recent in the CCT gene family. Our analyses show significant differences in the rates of evolution of archaeal chaperonins compared with the eukaryotic CCTs, as well as among the different CCT subunits themselves. We discuss these results in light of current views on the origin, evolution, and function of CCT complexes. (+info)
Dependence of Trichomonas vaginalis upon polyamine backconversion.
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Trichomonas vaginalis grown for 16 h in the presence of [(14)C]spermine formed a high intracellular pool of [(14)C]spermidine and a small but detectable pool of [(14)C]putrescine. When [(3)H]putrescine was added to the growth medium, a large intracellular pool of [(3)H]putrescine was found, but it was not further metabolized, confirming previous studies suggesting the absence of a forward-directed polyamine synthetic pathway in T. vaginalis. Spermidine:spermineN:(1)-acetyltransferase (SSAT) and polyamine oxidase enzyme activities were detected which collectively converted spermine to spermidine. Polyamine oxidase was localized in the hydrogenosome-enriched fraction, whereas SSAT was found predominantly in the cytosolic fraction. In the presence of saturating substrate, the trichomonad SSAT had an activity of 0. 39+/-0.09 nmol min(-1) (mg protein)(-1) (the mean of five analyses) and an apparent K:(m) for spermine of 1.7 microM. The enzyme was competitively inhibited by di(ethyl)norspermine with a K:(i) of 28 microM. Growth studies indicated that 50 microM di(ethyl)norspermine caused a 68% and 84% reduction in the intracellular concentrations of spermidine and spermine, respectively. The trichomonad polyamine oxidase required FAD as a cofactor and had an apparent K:(m) of 6.0 microM for N(1)-acetylspermine. The potential of bis(alkyl) polyamine analogues as antitrichomonad agents is discussed. (+info)
Trichomonas vaginalis epidemiology: parameterising and analysing a model of treatment interventions.
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BACKGROUND: Trichomonas vaginalis, which affects at least 170 million individuals globally, may increase the risk of transmission of HIV and predispose pregnant women to premature rupture of membranes and early labour. OBJECTIVE: To more clearly define the epidemiology of trichomoniasis and to develop a mathematical model of disease transmission dynamics in order to explore various treatment strategies. DESIGN: A deterministic model of trichomoniasis was constructed. Parameter values were set to fit the model to known endemic prevalence levels of Trichomonas vaginalis. Two treatment interventions ("screening" and "syndromic management") were simulated. RESULTS: The age specific prevalence of the disease was seen to differ from other STDs in a number of studies. Parameter fitting indicates that the average duration of infection in women is at least 3-5 years and approximately 4 months for men. "Syndromic management" (that is, treating only those with symptoms of disease) had minimal effect upon the endemic prevalence of disease even at high levels of coverage. "Screening" (that is, identification of individuals with both symptomatic and asymptomatic infection) was shown to be the most efficient method of control, but was sensitive to the screening interval. CONCLUSIONS: The control of trichomoniasis seems to have been a success in developed countries because of the regular access to health care, whereas it has remained endemic in many developing countries where control may only be possible by regular screening and treatment. However, without a large investment in services, success in controlling trichomoniasis is likely to be transitory. (+info)
Iron hydrogenases and the evolution of anaerobic eukaryotes.
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Hydrogenases, oxygen-sensitive enzymes that can make hydrogen gas, are key to the function of hydrogen-producing organelles (hydrogenosomes), which occur in anaerobic protozoa scattered throughout the eukaryotic tree. Hydrogenases also play a central role in the hydrogen and syntrophic hypotheses for eukaryogenesis. Here, we show that sequences related to iron-only hydrogenases ([Fe] hydrogenases) are more widely distributed among eukaryotes than reports of hydrogen production have suggested. Genes encoding small proteins which contain conserved structural features unique to [Fe] hydrogenases were identified on all well-surveyed aerobic eukaryote genomes. Longer sequences encoding [Fe] hydrogenases also occur in the anaerobic eukaryotes Entamoeba histolytica and Spironucleus barkhanus, both of which lack hydrogenosomes. We also identified a new [Fe] hydrogenase sequence from Trichomonas vaginalis, bringing the total of [Fe] hydrogenases reported for this organism to three, all of which may function within its hydrogenosomes. Phylogenetic analysis and hypothesis testing using likelihood ratio tests and parametric bootstrapping suggest that the [Fe] hydrogenases in anaerobic eukaryotes are not monophyletic. Iron-only hydrogenases from Entamoeba, Spironucleus, and Trichomonas are plausibly monophyletic, consistent with the hypothesis that a gene for [Fe] hydrogenase was already present on the genome of the common, perhaps also anaerobic, ancestor of these phylogenetically distinct eukaryotes. Trees where the [Fe] hydrogenase from the hydrogenosomal ciliate Nyctotherus was constrained to be monophyletic with the other eukaryote sequences were rejected using a likelihood ratio test of monophyly. In most analyses, the Nyctotherus sequence formed a sister group with a [Fe] hydrogenase on the genome of the eubacterium Desulfovibrio vulgaris. Thus, it is possible that Nyctotherus obtained its hydrogenosomal [Fe] hydrogenase from a different source from Trichomonas for its hydrogenosomes. We find no support for the hypothesis that components of the Nyctotherus [Fe] hydrogenase fusion protein derive from the mitochondrial respiratory chain. (+info)