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Resistance mechanisms of Trichomonas vaginalis to metronidazole are still not well understood. It has been shown that Mycoplasma hominis has the ability to establish an endosymbiotic relationship with T. vaginalis. This study investigated the association between T. vaginalis and M. hominis symbiosis in relation to metronidazole resistance. This study included 362 pregnant women from the King Edward VIII hospital in South Africa. The women provided self-collected vaginal swabs for the diagnosis of T. vaginalis by culture. Metronidazole susceptibility using the broth-microdilution assay was performed. Detection of the 16S rRNA from M. hominis using T. vaginalis genomic DNA as the template was performed. All statistical analysis was conducted in R statistical computing software. A total of 21 culture positive isolates were obtained resulting in a prevalence of 5.8% for T. vaginalis in the study population. Under anaerobic incubation, 52.4% (11/21) of the isolates were susceptible to metronidazole (MIC ≤ 1 μg/ml). Intermediate resistance (MIC of 2 μg/ml) and full resistance (4 μg/ml) was observed in 38.1% (8/21) and 9.5% (2/21) of the isolates, respectively. The majority of the isolates 95% (19/20) were susceptible to metronidazole under aerobic conditions. Only one isolate had a MIC of 50 μg/ml. M. hominis was shown to be present in 85.7% (18/21) of the T. vaginalis isolates. However, there was no significant association between metronidazole susceptibility and T. vaginalis-M. hominis symbiosis. This study provides evidence of emerging metronidazole resistance in T. vaginalis. However, these resistance profiles were not associated with M. hominis symbiosis.

The flagellated protozoon Trichomonas vaginalis, responsible for trichomoniasis, can establish a symbiotic relationship with the bacterium Mycoplasma hominis and can harbor double-stranded RNA Trichomonasvirus (TVV). In this study, we investigated by real-time PCR the prevalence of the four TVVs and of M. hominis among 48 T. vaginalis strains isolated in Italy, and we evaluated a possible association with metronidazole resistance. Fifty percent of the analyzed trichomonad strains tested positive for at least one TVV T. vaginalis, with TVV2 being the most prevalent, followed by TVV1 and TVV3. Two T. vaginalis strains were infected by TVV4, detected in Europe for the first time. Interestingly, we found more than one TVV species in 75% of positive trichomonad strains. M. hominis was present in 81.25% of T. vaginalis isolates tested, and no statistically significant association was observed with the infection by any TVV. Metronidazole sensitivity of T. vaginalis isolates was evaluated in vitro, and no correlation was observed between minimal lethal concentration and the presence of TVVs. This is the first report on TVV infection of T. vaginalis in Italy. Even if no association of TVV positive isolates with the presence of the symbiont M. hominis or with metronidazole resistance was observed, further studies are needed to shed light on the effective role of infecting microorganisms on the pathophysiology of T. vaginalis.

Mycoplasma lipoproteins play a relevant role in pathogenicity and directly interact with the host immune system. Among human mycoplasmas, Mycoplasma hominis is described as a commensal bacterium that can be associated with a number of genital and extragenital conditions. Mechanisms of M. hominis pathogenicity are still largely obscure, and only a limited number of proteins have been associated with virulence. The current study focused on investigating the role of MHO_0730 as a virulence factor and demonstrated that MHO_0730 is a surface lipoprotein, potentially expressed in vivo during natural infection, acting both as a nuclease with its amino acidic portion and as a potent inducer of Neutrophil extracellular trapsosis with its N-terminal lipid moiety. Evidence for M. hominis neutrophil extracellular trap escape is also presented. Results highlight the relevance of MHO_0730 in promoting infection and modulation and evasion of innate immunity and provide additional knowledge on M. hominis virulence and survival in the host., (© The Author(s) 2019. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

Trichomonas vaginalis is an anaerobic protist, responsible for the most prevalent non-viral sexually transmitted infection in humans. One of the most intriguing aspects of T. vaginalis pathobiology is the complex relationship with intracellular microbial symbionts: a group of dsRNA viruses belonging to family of Totiviridae (T. vaginalis virus), and eubacteria belonging to the Mycoplasma genus, in particular Mycoplasma hominis. Both microorganisms seem to strongly influence the lifestyle of T. vaginalis, suggesting a role of the symbiosis in the high variability of clinical presentation and sequelae during trichomoniasis. In the last few years many aspects of this unique symbiotic relationship have been investigated: M. hominis resides and replicates in the protozoan cell, and T. vaginalis is able to pass the bacterial infection to both mycoplasma-free protozoan isolates and human epithelial cells; M. hominis synergistically upregulates the proinflammatory response of human monocytes to T. vaginalis. Furthermore, the influence of M. hominis over T. vaginalis metabolism and physiology has been characterized. The identification of a novel species belonging to the class of Mollicutes (Candidatus Mycoplasma girerdii) exclusively associated to T. vaginalis opens new perspectives in the research of the complex series of events taking place in the multifaceted world of the vaginal microbiota, both under normal and pathological conditions.

Objectives: Mycoplasma hominis, a genetically heterogeneous, cell-wall-less bacterium, is able to live in symbiosis with the protozoan parasite Trichomonas vaginalis. Whilst the impact of this symbiosis on T. vaginalis has been investigated to a certain extent, less light has been shed on the influence on M. hominis., Methods: An in vitro minimum inhibitory concentration (MIC) study of the antimicrobial susceptibility of three clinical M. hominis isolates (V475, AKH136 and MhSS10) to clindamycin, moxifloxacin, ciprofloxacin and gentamicin was performed in dependence on symbiosis with T. vaginalis strain IR78., Results: Passaging of M. hominis through T. vaginalis led to an increase in MICs to all drugs investigated in M. hominis V475 and M. hominis MhSS10 (apart from gentamicin). Shifts from intermediate to resistant (MhSS10 for ciprofloxacin) and from susceptible to intermediate-resistant (V475 for gentamicin; P=0.015) were observed. Moreover, initial susceptibility of V475 to moxifloxacin (MIC=1.35μg/mL) was statistically significantly reduced (MIC=2.5μg/mL) following T. vaginalis passage concomitantly with mutations in the quinolone resistance-determining regions (QRDRs) of gyrA (S153L) and parC (E195G and K144R). In contrast, the susceptibility of M. hominis isolate AKH136 to all drugs investigated increased after passaging., Conclusions: These findings suggest that symbiosis with T. vaginalis has an enhancing effect on selected antimicrobial resistances of distinct M. hominis isolates., (Copyright © 2018 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Mycoplasma hominis is a human urogenital pathogen involved in gynaecological, neonatal and extra-genital infections. However, no versatile genetic tools are currently available to study the pathogenicity of this bacterium. Targeting-Induced Local Lesions IN Genomes (TILLING) is a reverse-genetic method that combines point mutations induced by chemical mutagenesis with a DNA screening technique. We used ethyl methanesulfonate (EMS) that introduces C-G to T-A transition mutations to generate a library of M. hominis mutants. As a proof of concept, mutagenized organisms were screened for mutations in two target genes previously associated with the mycoplasma pathogenicity, the vaa gene encoding an adhesin lipoprotein and the oppA gene encoding the main ectoATPase of the bacterium. The resulting mutants were evaluated using functional assays, an adhesion to HeLa cell assay for vaa-mutants and an ATPase activity test for oppA-mutants., Results: A 1200-clone library was generated by exposing M. hominis PG21 to 9 mg/mL EMS for 3 h. To identify mutants of interest, targeted gene fragments were amplified, heat-denatured, slowly reannealed and digested with the mismatch-specific endonuclease ENDO1. If multiple alleles were present in the PCR amplicons, these alleles formed heteroduplexes during reannealing that were specifically cleaved by ENDO1 at mismatching positions. A total of four vaa-mutants and two oppA-mutants harbouring missense mutations were obtained and fully sequenced. Zero to eight additional mutations were identified in the genomes of each mutant. The vaa-mutants were tested for adhesion to immobilized HeLa cells but their adhesion was not significantly different from the adhesion of M. hominis PG21. One of the two oppA-mutants that were tested for ATPase activity presented a higher affinity for its ATP substrate than the parental strain., Conclusion: For the first time, we demonstrated that M. hominis gene-targeted mutants could be successfully obtained using this TILLING strategy. In the absence of robust genetic tools for studying M. hominis, the TILLING strategy that can target any gene of the genome could help to elucidate gene functions and to better understand the pathogenesis of this human pathogenic species.

Cancer was initially considered a genetic disease. However, recent studies have revealed the connection between bacterial infections and growth of different types of cancer. The enteroinvasive strain of Mycoplasma hominis alters the normal behavior of host cells that may result in the growth of prostate cancer. The role of M. hominis in the growth and development of prostate cancer still remains unclear. The infection may regulate several factors that influence prostate cancer growth in susceptible individuals. The aim of this study was to predict M. hominis proteins targeted into the endoplasmic reticulum (ER) of the host cell, and their potential role in the induction of prostate cancer. From the whole proteome of M. hominis , 19 proteins were predicted to be targeted into the ER of host cells. The results of our study predict that several proteins of M. hominis may be targeted to the host cell ER, and possibly alter the normal pattern of protein folding. These predicted proteins can modify the normal function of the host cell. Thus, the intercellular infection of M. hominis in host cells may serve as a potential factor in prostate cancer etiology.

In Europe, up to 90% of isolated Trichomonas vaginalis strains are naturally infected with Mycoplasma hominis, a facultative pathogen of the human genital tract. The consequences of this endosymbiosis are not yet well understood. The aim of the current study was to evaluate the impact of natural and artificial infections with M. hominis on the RNA expression levels of metronidazole susceptibility-associated genes of T. vaginalis. Three T. vaginalis strains (TVSS10-, TVSS25-, G3) without M. hominis, as well as the same strains naturally (TVSS10+, TVSS25+) and artificially (G3-MhSS25, TVSS25-MhSS25) infected with M. hominis, were investigated for their expression profiles of three genes associated with metronidazole resistance (ferredoxin, flavin reductase 1 and pyruvate:ferredoxin oxidoreductase). The minimal inhibitory concentrations (MICs) of metronidazole were evaluated for all combinations and the respective M. hominis-free T. vaginalis strains were used as controls. The sole presence of M. hominis led to a down-regulation of metronidazole susceptibility-associated genes in all T. vaginalis strains tested. Interestingly, the effect was more prominent in the artificial symbioses. Moreover, a twofold enhancement of metronidazole tolerability was observed in three infected T. vaginalis strains, compared to the respective strains without M. hominis. In conclusion, M. hominis had an impact on gene expression in all T. vaginalis strains and on metronidazole MIC in all but one strain tested.

We report the first case of Mycoplasma hominis periaortic abscess after heart-lung transplantation. The absence of sternal wound infection delayed the diagnosis, but the patient successfully recovered with debridement surgeries and long-term antibiotic therapy. Owing to the difficulty in detection and the intrinsic resistance to beta-lactams, M. hominis infections are prone to being misdiagnosed and undertreated. M. hominis should be suspected in cases where conventional microbiological identification and treatment approaches fail., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Although the idea of bacteria causing different types of cancer has exploded about century ago, the potential mechanisms of carcinogenesis is still not well established. Many reports showed the involvement of M. hominis in the development of prostate cancer, however, mechanistic approach for growth and development of prostate cancer has been poorly understood. In the current study, we predicted M. hominis proteins targeting in the mitochondria and cytoplasm of host cells and their implication in prostate cancer. A total of 77 and 320 proteins from M. hominis proteome were predicted to target in the mitochondria and cytoplasm of host cells respectively. In particular, various targeted proteins may interfere with normal growth behaviour of host cells, thereby altering the decision of programmed cell death. Furthermore, we investigated possible mechanisms of the mitochondrial and cytoplasmic targeted proteins of M. hominis in etiology of prostate cancer by screening the whole proteome.

Mycoplasma hominis (M. hominis) and Ureaplasma urealyticum (U. urealyticum) are important opportunistic pathogens that cause urogenital infections and complicate pregnancy. The aim of this study was to investigate the prevalence, effects on pregnancy outcomes, and antimicrobial susceptibilities of M. hominis and U. urealyticum. We tested vaginal swabs obtained from 1035 pregnant women for the presence of genital mycoplasmas between June 2009 and May 2014. The laboratory and clinical aspects of genital mycoplasmas infection were reviewed retrospectively, and the identification and antimicrobial susceptibility of genital mycoplasmas were determined using the Mycoplasma IST-2 kit. A total of 571 instances of M. hominis and/or U. urealyticum were detected. Of them, M. hominis was detected in two specimens, whereas U. urealyticum was detected in 472 specimens. The remaining 97 specimens were positive for both M. hominis and U. urealyticum. Preterm deliveries were frequently observed in cases of mixed infection of M. hominis and U. urealyticum, and instances of preterm premature rupture of membrane were often found in cases of U. urealyticum. The rates of non-susceptible isolates to erythromycin, empirical agents for pregnant women, showed increasing trends. In conclusion, the prevalence of M. hominis and/or U. urealyticum infections in pregnant women is high, and the resistance rate of antimicrobial agents tends to increase. Therefore, to maintain a safe pregnancy, it is important to identify the isolates and use appropriate empirical antibiotics immediately.

Gardnerella vaginalis plays an important role in bacterial vaginosis (BV,) while the role of genital Mollicutes is less obvious. The diagnosis of BV by use of the current Gram stain Nugent score is also suboptimal for defining the role of Mollicutes that lack a cell wall. Since bacterial load and diversity is an important prerequisite for BV, real-time quantitative polymerase chain reaction (qPCR) assays enable these to be assessed. The purpose of this study was to define the role of genital Mollicutes and potential patterns of synergy with G. vaginalis in women with BV. Vaginal swabs from 130 women categorised by Nugent score as BV (n = 28), intermediate (n = 22) and non-BV (n = 80) were tested against four qPCR TaqMan assays targeting G. vaginalis, Mycoplasma hominis, M. genitalium, Ureaplasma urealyticum and U. parvum. Statistical analyses were used to compare bacterial prevalence and load between the three groups of women. Mycoplasma hominis and G. vaginalis co-infection was significantly more common in BV (60.7 %) compared to intermediate (36.4 %) and non-BV (8.8 %) Nugent scores (p < 0.001). Significantly higher loads of M. hominis (p = 0.001) and G. vaginalis (p < 0.001) were detected in women with BV and the respective loads in M. hominis and G. vaginalis co-infections displayed a significant positive correlation (p < 0.001; r = 0.60). No significant associations were seen with the other Mollicutes. The findings strengthen the evidence of a role for M. hominis in BV and a potential synergy with G. vaginalis. This synergy could be an important trigger of the condition and sexual contact the conduit for the transmission of an otherwise commensal bacterium that could initiate it.

Objectives: Trichomonas vaginalis is the causative agent of trichomoniasis, one of the most common sexually transmitted diseases worldwide. In recent years we have described the symbiotic relationship between T vaginalis and Mycoplasma hominis. How this biological association might affect the pathogenicity of one or both the microorganisms is still unknown. Since local inflammation is thought to play a central role in T vaginalis infection, we investigated the in vitro response of human macrophages to naturally mycoplasma-free T vaginalis, as compared to a mycoplasma-infected trichomonad isolate., Methods: THP-1 cells were stimulated with two isogenic T vaginalis isolates, one naturally mycoplasma-free and one stably associated with M hominis, and secreted cytokines measured by ELISA. Nuclear factor κB (NFκB) involvement in THP-1 response to T vaginalis and M hominis was evaluated by means of a reporter system based on detection of alkaline phosphatase activity., Results: We found that the presence of M hominis upregulates the expression of a panel of proinflammatory cytokines in a synergistic fashion. We also found that the upregulation of the proinflammatory response by THP-1 cells involves the transcription factor NFκB., Conclusions: These findings suggest that the presence of M hominis in T vaginalis isolates might play a key role in inflammation during trichomoniasis, thus affecting the severity of the disease. The synergistic upregulation of the macrophage proinflammatory response might also affect some important clinical conditions associated with T vaginalis infection, such as the increased risk of acquiring cervical cancer or HIV, which are thought to be affected by the inflammatory milieu during trichomoniasis.

Here we examined the involvement of Mycoplasma hominis in the formation of biofilms by uropathogenic Escherichia coli (UPEC) strain CFT073. Initially, we thought that M. hominis does not affect the fitness of UPEC, including the growth and production of signaling molecules, such as autoinducer-2 and indole. We found, however, that the presence of M. hominis significantly decreased the degree of biofilm formation by UPEC CFT073 (approximately a 60% reduction for 10(5) ccu/mL of M. hominis as compared with UPEC alone). We also found that it had a slight effect in inhibiting the attachment and cytotoxicity of UPEC CFT073. These findings are specific to these UPEC strains rather than to enterohemorrhagic E. coli (EHEC) strains, found in normal intestinal flora. In addition, we performed whole-transcriptome profiling and quantitative real-time polymerase chain reaction (qRT-PCR) analysis. This indicated that the PhoPQ system and the anti-termination protein (encoded by ybcQ) were involved in the reduction of biofilm formation by M. hominis (corroborated by qRT-PCR). Furthermore, our results indicate that M. hominis raises the degree of transcription of toxin genes, including hha and pasT. Hence, we suggest a possible role of M. hominis in affecting the formation of biofilms by UPEC in the urinary tract.

Background: In Mycoplasma hominis, a facultative human pathogen of the human genital tract, OppA, the substrate-binding domain of the oligopeptide permease, is a multifunctional protein involved in nutrition uptake, cytoadhesion and hydrolysis of extracellular ATP., Results: To map the function-related protein regions the ATPase activity and adhesive behavior of OppA mutants were analyzed. Mutations of the Walker BA motifs resulted in an inhibition of up to 8% of the OppA ATPase activity, whereas deletion of the N-terminal CS1 or the CS2 region, structural motifs that are conserved in bacterial OppA proteins, reduced ATPase activity to 60% and deletion of CS3, the third conserved region adjacent to the Walker B motif led to a reduction to 42% ATPase activity. Interestingly, adhesion of the OppA mutants to immobilized HeLa cells demonstrated that two distal regions are mainly involved in adherence of OppA: the CS1 region, deletion of which led to 35% of the cytoadhesion, and the Walker BA with the adjacent upstream region CS3, deletion of which led to 25% of the cytoadhesion. The influence of the ATPase activity on the adherence of M. hominis to HeLa cells was confirmed by the use of ATPase inhibitors which reduced mycoplasmal cytoadhesion to 50%., Conclusions: These findings suggest that the OppA-mediated cytoadherence of Mycoplasma hominis depends on both, the topology of the neighbouring CS1 and ATPase domain regions and the functionality of the ecto-ATPase activity in addition.

We isolated 30 Trichomonas vaginalis for the PCR detection from the gynecological outpatients in the Affiliated Hospital of Zhengzhou University using the specific 16s rDNA primers of Mycoplasma hominis. The results showed that there were 25 cases of Mycoplasma hominis infection, with the infection rate of 83.33%. This gave a clew that the symbiosis of Trichomonas vaginalis with Mycoplasma hominis may be of certain generality in China. We sequenced the ferredoxin gene of 10 Trichomonas vaginalis where 5 Mycoplasma hominis were positive and five negative, and found that the ferredoxin (Fd) gene of the 10 Trichomonas vaginalis were exactly the same. But compared to the genes in the GenBank, a comparative analysis of the gene revealed that there were 3 more ctg bases at the 200th position of encoding leucine, but this did not lead to changes in reading frame. The gene homology was 99%.

Problem: We compared the frequency of intra-amniotic infection in preterm labor (PL) with women not in labor, and correlated infection with amniotic fluid (AF) cytokines. Detailed identification of species, especially mycoplasmata, was tried to improve our understanding of the pathogenesis of PL., Method of Study: AF from 20 women with PL and 20 controls were evaluated. Infection was detected by PCR for Mycoplasma hominis, Ureaplasma urealyticum and 16S rRNA bacterial gene, which was cloned and sequenced for bacterial identification. Interleukin (IL)-1β, IL-6, IL-8 and tumor necrosis factor (TNF)-α levels were measured by ELISA., Results: Frequency of intra-amniotic infection is higher in PL (40.0%). Sequencing-based method identified Bacteroides fragilis, Prevotella bivia and Leptotrichia amnionii, in addition to Mycoplasma species detected by PCR. AF infection correlated with increased IL-1β and IL-6 levels., Conclusion: The frequency of intra-amniotic infection, especially M. hominis, in PL women who delivered with 7 days, is high and correlates with high IL-1β and IL-6 levels, but not IL-8., (© 2011 John Wiley & Sons A/S.)

The origin of chronic inflammation preceding the development of prostate cancer (PCa) remains unknown. We investigated possible involvement of mycoplasma infection in PCa by screening prostate biopsies from two groups of Russian men undergoing PCa diagnosis. M. hominis was detected by standard PCR in 15% of the 125 patients in the first group and by quantitative real-time PCR in 37.4% of the 123 men in the second group. In both groups, stratification of patients according to diagnosis showed that M. hominis was present at three times higher frequency in patients with PCa than in those with benign prostatic hyperplasia. No M. hominis was detected in the prostates of 27 men without detectable prostate disease. In addition, PCa-positive men had higher titers of antibodies against M. hominis and average PSA levels were higher in M. hominis-positive men. These data, together with previous observations linking mycoplasma infection with cell transformation, genomic instability and resistance to apoptosis, suggest that M. hominis infection may be involved in PCa development and may, therefore, be a potential PCa marker and/or target for improved prevention and treatment of this disease.

We report a case of neonatal subdural empyema caused by Mycoplasma hominis. The infant sustained severe birth-related eye injury and subsequently developed multiple subdural empyemata. This report illustrates the clinical presentation of this unusual infection and the crucial role of neurosurgical intervention and specific antimicrobial therapy for its successful management.

Objective: Microbial biofilms are communities of sessile microorganisms formed by cells that are attached irreversibly to a substratum or interface or to each other and embedded in a hydrated matrix of extracellular polymeric substances. Microbial biofilms have been implicated in >80% of human infections such as periodontitis, urethritis, endocarditis, and device-associated infections. Thus far, intraamniotic infection has been attributed to planktonic (free-floating) bacteria. A case is presented in which "amniotic fluid sludge" was found to contain microbial biofilms. This represents the first report of a microbial biofilm in the amniotic cavity., Study Design: "Amniotic fluid sludge" was detected by transvaginal sonography and retrieved by transvaginal amniotomy. Bacteria were identified with scanning electron microscopy and fluorescence in situ hybridization for conserved regions of the microbial genome; the exopolymeric matrix was identified by histochemistry by the wheat germ agglutinin lectin method. The structure of the biofilm was imaged with confocal laser scanning microscopy., Results: "Amniotic fluid sludge" was imaged with scanning electron microscopy, which allowed the identification of bacteria embedded in an amorphous material and inflammatory cells. Bacteria were demonstrated with fluorescent in situ hybridization using a eubacteria probe. Extracellular matrix was identified with the wheat germ agglutinin lectin stain. Confocal microscopy allowed 3-dimensional visualization of the microbial biofilm., Conclusion: Microbial biofilms have been identified in a case of intraamniotic infection with "amniotic fluid sludge."

The parasite Trichomonas vaginalis causes one of the most common non-viral sexually transmitted infections in humans. The coexistence of different sexually transmitted diseases in the same individual is very common, such as vaginal infections by T. vaginalis in association with Mycoplasma fermentans or Mycoplasma hominis. However, the consequences and behavior of mycoplasma during trichomonad infections are virtually unknown. This study was undertaken to elucidate whether mycoplasmas enter and leave trichomonad cells and if so how. M. hominis was analyzed in different trichomonad isolates and the process of internalization and the pathway within the parasite was studied. Parasites naturally and experimentally infected with mycoplasmas were used and transmission electron microscopy, cytochemistry and PCR analyses were performed. The results show that: (1) M. hominis enters T. vaginalis cells by endocytosis; (2) some mycoplasmas use a terminal polar tip as anchor to the trichomonad plasma membrane; (3) some trichomonad isolates are able to digest mycoplasmas, mainly when the trichomonads are experimentally infected; (4) some fresh virulent isolates are able to maintain mycoplasmas as cohabitants in the cell's interior; (5) some mycoplasmas are able to escape from the vacuole to the trichomonad cytosol, and trichomonad plasma membrane budding suggested that mycoplasmas could leave the parasite cell.

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.

Mollicutes are the smallest and simplest self-replicating microorganisms. Despite the minimal genome and apparent lack of complexity, mycoplasmas show a high degree of adaptation to the most diverse environments. Mycoplasma hominis is a human sexually transmitted mycoplasma which is able to establish a biological association with Trichomonas vaginalis, a pathogenic flagellated protist. M. hominis and T. vaginalis share the same specific natural niche, the human genitourinary tract. Symbiotic relationships between unicellular eukaryotes and bacteria are well known and have been extensively studied, providing interesting insights into the biology of one or both the symbionts. The relationship between T. vaginalis and M. hominis is unique in that it was the first described association of two obligated human parasites. Several aspects of this relationship have been investigated, showing how the trichomonad may be viewed not only as a new niche for M. hominis, but also as a "Trojan horse" for the transmission of the bacterial infection to the human host.

The effect of cocultivation of eukaryotic HeLa cells and Mycoplasma hominis mycoplasma on the resistance of the latter to fluoroquinolones (ciprofloxacin) was examined. It was shown that cocultivation of the M. homonis and HeLa cells during 24 h with subsequent addition of ciprofloxacin resulted in an increase of the mircoplasma resistance to this antimicrobial agent. In the M. hominis cells cultivated in the presence of HeLa cells and the increasing concentration of ciprofloxacin mutations in the parC gene were observed only at low concentrations of the antimicrobial agent, while mutations in the gyrA gene were never detected. A gradual elevation of ciprofloxacin concentration up to 10 micrograms/ml resulted in the reversion of the parC mutations in mycoplasmas. Mycoplasma cells resistant to high flouroquinolone concentrations and isolated after cocultivation with the HeLa cells were characterized by the wild-type genotype in respect of the gyrA and parC genes. It was shown for the first time that infection of HeLa cells resulted in the appearance of genome rearrangements in M. hominis cells.

As the number of pathogenic microbial strains resistant to different antibiotics increases, amphipathic peptides with antimicrobial activity are promising agents for the therapy of infectious diseases. This work deals with the effect of an amphipathic antimicrobial peptide, melittin, expressed within recombinant plasmid vectors, on infection with urogenital pathogens Chlamydia trachomatis and Mycoplasma hominis in HeLa cell culture. Recombinant plasmid constructs with the melittin gene under the control of the tetracycline-responsive promoter of human cytomegalovirus were obtained. We showed inhibition of C. trachomatis and M. hominis infection after the introduction of recombinant plasmid vectors expressing the melittin gene into the infected cell culture.

Vaginal infections by Trichomonas vaginalis and Mycoplasma hominis have been shown to be associated. Since M. hominis and Ureaplasma urealyticum are similar pathogens, both belonging to the class of the mycoplasmata, we describe here a molecular study into the interdependence of U. urealyticum and T. vaginalis during infection. Susceptibility towards infection by U. urealyticum depends on genetic polymorphism in the interleukin-1 receptor antagonist (IL-1RA) gene. Now, we defined the relation between IL-1RA genotypes and infection by M. hominis and T. vaginalis. Finally, we also developed a restriction fragment length polymorphism (RFLP) tool for mapping variation in the T. vaginalis AP33 adhesin in order to define putative associations between parasite subtype and mycoplasmata or host. Studies using crudepellets from T. vaginalis culture broth clearly confirm the association between T. vaginalis and M. hominis infection. The association between IL-1RA genotype 2,2 and lack of U. urealyticum infection is corroborated as well. U. urealyticum infection and infection by T. vaginalis are independent. Furthermore, T. vaginalis and M. hominis infection are not depending on IL-1RA genotypes. Interestingly, one of the three AP33 RFLP types identified appeared to be associated with the absence of U. urealyticum infection. In conclusion, the complex interaction between bacterial and parasitic pathogens and the infected host is determined by genetic characteristics of host and microorganisms involved.

Background: Very small preterm infants who have genital mycoplasmas isolated from the trachea are at increased risk to develop bronchopulmonary dysplasia (BPD). The early stages of BPD are characterized by inflammation. Recruitment and activation of mononuclear cells in response to mycoplasmas may be important in the early stage of the disease. Lung epithelial cell production of monocyte chemoattractant protein-1 (MCP-1), a protein that attracts and activates mononuclear cells, could be critical in the regulation of mononuclear cell migration to the lung., Methods: We examined the potential of Mycoplasma hominis (Mh) to induce MCP-1 gene expression and protein production in A549 cells, a pulmonary epithelial cell line with characteristics of type II cells., Results: Live or heat-inactivated Mh induces MCP-1 mRNA and protein in a dose- and time-dependent manner. Stimulation of MCP-1 by Mh was not inhibited by 50 micrograms/mL of polymyxin B, interleukin (IL)-1ra, or neutralizing antibodies to IL-1 alpha, IL-1 beta, or tumor necrosis factor alpha (TNF-alpha). IL-1 alpha, IL-1 beta, and TNF-alpha were not detected in conditioned media of Mh-stimulated A549 cells., Conclusions: These data suggest that Mh may participate in the inflammatory component of BPD by directly inducing epithelial cell production of cytokines that recruit and activate mononuclear cells.

A characteristic feature of the mycoplasmas is the presence of variable surface proteins which may play an important role in the adaptation of the cell-wall-less organisms to their host environments. In addition, this antigen variation may be an important pathogenic property of the organism. The ubiquity of the gene encoding P50, an adhesin of Mycoplasma hominis FBG, and its transcription were analysed in different isolates of M. hominis. The p50 gene was present in all isolates tested. Based on Southern blot analysis and sequencing of the gene, the isolates could be classified into one of three distinct groups. Within two groups specific truncations of the p50 gene occurred. The reduction of the gene size was confirmed in Northern blot analysis of representative isolates from each group, with a decrease in transcript length from 1.6 kb in group G-1 down to 0.76 kb in group G-3. In addition to truncation, a coincidental duplication of some gene segments was detected. This work has provided evidence for the genetic basis of a further variation in the M. hominis P50 adhesin.

Objective: Our goal was to evaluate the effect of Escherichia coli, Bacteroides fragilis, Mycoplasma hominis, and Staphylococcus aureus on cytokine and prostaglandin production by amnion cells in vitro., Study Design: Amnion cells were obtained from women undergoing elective cesarean section before the onset of labor and cultured in a primary cell culture. Confluent amnion cells were incubated with heat-inactivated bacteria in different concentrations (10(1) to 10(6) colony-forming units/ml) for 48 hours. Samples for quantification of interleukin-1 beta, interleukin-6, interleukin-8, tumor necrosis factor-alpha, and prostaglandin E2 were collected at 6, 12, 24, and 48 hours., Results: Under basal conditions, minor amounts of interleukin-6 and interleukin-8 were detectable. Incubation of amnion cells with E. coli enhanced the secretion of interleukin-8 and also induced an transient increase of prostaglandin E2 in a dose-dependent manner. B. fragilis produced an increase in the secretion of interleukin-6 and interleukin-8. M. hominis and S. aureus did not cause an increase in either interleukin-6, interleukin-8, or prostaglandin E2., Conclusion: The gram-negative bacteria E. coli and B. fragilis stimulated interleukin-6 and interleukin-8 to a greater degree than the other bacteria investigated in this study. This finding may be of clinical interest in the onset of preterm birth.