Colonization of the respiratory and genital tracts of female mice with Mycoplasma pneumoniae and protection afforded to the genital tract by prior respiratory colonization

Mycoplasma pneumoniae, strain MY 12763, colonized the throats of 6 BALB/c female mice for at least 18 days after intranasal inoculation. The same strain colonized, in high titre, the vagina of 9 of 10 progesterone-treated BALB/c mice for at least 35 days, but none of 10 oestradiol-treated mice. Mice were less susceptible to genital-tract colonization with the multiple broth-passed FH strain of M. pneumoniae, only 3 of 10 becoming colonized. The 6 mice inoculated intranasally with strain MY 12763 were immune to genital-tract colonization with the same strain, whereas 10 mice without respiratory-tract colonization were susceptible. Protection of the genital tract in this way was at least as effective as that afforded by previous genital-tract colonization. In a further experiment, 26 immunocompetent BALB/c mice colonized previously in the respiratory tract were resistant to vaginal colonization, whereas 20 BALB/c nude mice, similarly colonized in the respiratory tract, were susceptible in the vagina, illustrating the importance of cell-mediated immunity. The possible relevance of the findings to the human situation is discussed.     

Comparison of host responses after intranasal infection of guinea-pigs with Mycoplasma genitalium or with Mycoplasma pneumoniae

Guinea-pigs were infected intranasally with Mycoplasma genitalium or Mycoplasma pneumoniae. The lung lesions produced by the two mycoplasmas were comparable in extent and histological pattern. Sera of both animal groups taken 2 weeks after infection reacted strongly in the complement fixation test with the M. pneumoniae glycolipid extract. In an ELISA using the respective adherence proteins (P1-protein of M. pneumoniae and MgPa of M. genitalium), strong specific activity, but also considerable cross-reactions were found. Epitope analysis by using overlapping octapeptides of a P1-region immunologically active in human M. pneumoniae infections and of the corresponding MgPa-region revealed six common epitopes but also one M. genitalium and two M. pneumoniae specific determinants. For analysis of a possible pathogenicity of M. genitalium in the human respiratory tract species-specific tests have to be developed.     

Isolation of Mycoplasma pneumoniae from the human urogenital tract.

Mycoplasma pneumoniae is a common etiologic agent of lower respiratory tract infections in humans. However, it has been reported previously that the organism has occasionally been isolated from sites other than the oropharynx and respiratory tract. We report the isolation of 24 strains of M. pneumoniae from urogenital specimens obtained from 22 female patients. Most isolates were of cervical origin from patients attending several local gynecological clinics over a 2-year period. Strains were also isolated from the urethra of one of three healthy male sexual partners of female patients positive for the organism. Single serum specimens obtained from three female patients and three different male sexual partners showed antibody levels suggestive of either recent respiratory infection or genital tract colonization with M. pneumoniae. Although there is no apparent definitive explanation for the localized outbreak of the organism at these unusual sites, the possible transfer through sexual and/or orogenital contact remains the most likely mode of transmission. The occurrence of an organism with obvious pathogenicity for human epithelial tissue in the urogenital tract suggests such transfer could play a role in genital tract infection.     

The susceptibility of germ-free, oestradiol-treated, mice to Mycoplasma hominis

Conventionally reared female BALB/c mice, rendered susceptible to Mycoplasma hominis infection of the genital tract by treatment with oestradiol, have increased numbers of endogenous vaginal bacteria. The latter was reflected by the occurrence of bacterial growth in 95 (65.5%) of 145 cultures undertaken to isolate M. hominis from oestradiol-treated mice, but in only seven (4.8%) of 146 cultures from untreated animals. In addition, larger numbers of bacteria were seen in vaginal smears from oestradiol-treated mice than from untreated ones. Furthermore, abscesses developed in the genital region of 27 (17%) of 155 oestradiol-treated mice but in none of 50 that were untreated. However, such proliferation of the endogenous vaginal bacteria was not necessary for colonisation of the vagina by M. hominis. This was determined by showing that six germ-free, oestradiol-treated BALB/c mice given 2.5 x 10(5) ccu of M. hominis intravaginally became colonised vaginally for at least 14 days, with multiplication and spread of the organisms to the upper genital tract and elsewhere, whereas six similar untreated mice given the same inoculum remained uninfected.     

Shared epitopes between Mycoplasma pneumoniae major adhesin protein P1 and a 140-kilodalton protein of Mycoplasma genitalium

Previous serological data have demonstrated cross-reactive antigens between two pathogenic species of mycoplasmas, M. pneumoniae and M. genitalium. Preliminary analysis of sera and monoclonal antibodies (MAbs) to protein antigens of these species showed an immunodominance of adhesin P1 (165 kilodaltons [kDa]) of M. pneumoniae in mice and hamsters and a 140-kDa protein of M. genitalium in mice and experimentally infected chimpanzees. To further characterize these two proteins, we assayed multiple anti-P1 and anti-140-kDa protein MAbs by enzyme-linked immunosorbent assay, immunoblot, and radioimmunoprecipitation techniques. The 140-kDa M. genitalium protein was shown to be surface accessible and insensitive to levels of trypsin which readily degrade protein P1. Peptide mapping was used to identify a unique class of MAbs which bound a cross-reactive molecule common to both the major adhesin protein P1 of M. pneumoniae and the 140-kDa protein of M. genitalium. MAbs generated against both M. pneumoniae and M. genitalium which were reactive with this determinant blocked M. pneumoniae attachment to chicken erythrocytes.     

Isolation and characterization of Mycoplasma genitalium strains from the human respiratory tract.

Mycoplasma genitalium, an organism first isolated from the urethras of two men with nongonococcal urethritis, has been found in throat specimens from military recruits participating in an inactivated Mycoplasma pneumoniae vaccine field trial in 1974-1975. Four of 16 preserved throat isolates, previously identified as strains of M. pneumoniae, have now been shown to be mixtures of M. pneumoniae and M. genitalium. Purification of these mixed mycoplasmas by selection of single colonies confirmed the presence of M. genitalium. Identification of M. genitalium was based upon the occurrence of a species-specific 140-kilodalton protein adhesin in these isolates and their serologic reactivity to an M. genitalium antiserum. The frequent occurrence of both M. pneumoniae and M. genitalium in a number of these throat specimens, in combination with their shared antigenic cross-reactivities, suggests the likelihood that M. genitalium strains are easily missed in the usual laboratory identification procedures. What role M. genitalium may play in human respiratory disease remains to be determined.     

Application of an indirect immunofluorescence test for detection of Mycoplasma pneumoniae in respiratory exudates.

We prepared polyclonal antibody specific to Mycoplasma pneumoniae and examined the conditions influencing the ability of an indirect immunofluorescence test to detect the specific antigen in respiratory exudates. The antibody did not cross-react with normal human serum or with respiratory exudates from 10 healthy persons. Cross-reactivity of the antibody with species of mycoplasmas other than M. genitalium was fully diminished when absorbed with horse serum and yeast extract, components of the culture medium. Though the absorbed antibody cross-reacted with M. genitalium, the titer was significantly lower than when tested against M. pneumoniae. Two types of antigen-specific fluorescence were observed in clinical specimens: one is large or small fluorescent granular aggregates found in mucus, and the other is fine fluorescent particles diffused on the entire surface of small epithelial cells. Throat smears from 49 patients with serologically confirmed M. pneumoniae infections were examined by our indirect immunofluorescence method. Positive results were obtained in 42 cases, many of which were positive before a rise in serum antibody titer could be demonstrated, indicating that the method is useful for a preliminary diagnosis at an early stage of the infection.     

A Mycoplasma genitalium protein resembling the Mycoplasma pneumoniae attachment protein.

In previous studies with hyperimmune rabbit sera and monoclonal antibodies against the P1 protein of Mycoplasma pneumoniae, we obtained evidence of a shared antigenic determinant with a single protein of Mycoplasma genitalium. Because of biologic and morphologic similarities between these two human Mycoplasma species, attempts were made to characterize this cross-reacting protein of M. genitalium (designated MgPa). The protein was surface exposed and had an estimated molecular size of 140 kilodaltons. Electron microscopy with monoclonal antibodies produced against either MgPa or P1 demonstrated that MgPa is located over the surface of the terminal structure of M. genitalium which is covered by a nap layer. These immunologic and morphologic findings suggest that the MgPa protein of M. genitalium could be the counterpart of the P1 protein of M. pneumoniae.     

DNA and protein sequence homologies between the adhesins of Mycoplasma genitalium and Mycoplasma pneumoniae.

Mycoplasma genitalium and Mycoplasma pneumoniae are morphologically and serologically related pathogens that colonize the human host. Their successful parasitism appears to be dependent on the product, an adhesin protein, of a gene that is carried by each of these mycoplasmas. Here we describe the cloning and determine the sequence of the structural gene for the putative adhesin of M. genitalium and compare its sequence to the counterpart P1 gene of M. pneumoniae. Regions of homology that were consistent with the observed serological cross-reactivity between these adhesins were detected at both DNA and protein levels. However, the degree of homology between these two genes and their products was much higher than anticipated. Interestingly, the A + T content of the M. genitalium adhesin gene was calculated as 60.1%, which is substantially higher tham that of the P1 gene (46.5%). Comparisons of codon usage between the two organisms revealed that M. genitalium preferentially used A- and T-rich codons. A total of 65% of positions 3 and 56% of positions 1 in M. genitalium codons were either A or T, whereas M. pneumoniae utilized A or T for positions 3 and 1 at a frequency of 40 and 47%, respectively. The biased choice of the A- and T-rich codons in M. genitalium could also account for the preferential use of A- and T-rich codons in conservative amino acid substitutions found in the M. genitalium adhesin. These facts suggest that M. genitalium might have evolved independently of other human mycoplasma species, including M. pneumoniae.     

Mycoplasma pneumoniae and Mycoplasma genitalium mixture in synovial fluid isolate.

A mycoplasma cultured from synovial fluid specimens from a patient with pneumonia and subsequent polyarthritis was identified initially as Mycoplasma pneumoniae. In retrospective studies, the culture was shown also to contain Mycoplasma genitalium. In this paper, the laboratory techniques employed in the identification and separation of the two species are presented, and evidence to implicate postinfectious autoimmunity is provided. An increasing number of reports of M. genitalium in human tissue sites and difficulties in isolation and identification of the organism in the clinical laboratory suggest the need for more extensive application of rapid and specific detection systems for both M. genitalium and M. pneumoniae in the clinical laboratory.     

Mycoplasma genitalium attaches to human spermatozoa

BACKGROUND: Mycoplasma genitalium causes urogenital diseases in men and women and is presumed to be sexually transmitted. We wanted to investigate whether spermatozoa could serve as vectors for M.genitalium in order to cause upper genital diseases in women. METHODS: By use of Nomarski light microscopy and transmission X-ray microscopy, the attachment of M.genitalium to spermatozoa was studied. Semen was incubated in vitro with M.genitalium. Purified, motile spermatozoa were examined for attachment of M.genitalium by immunofluorescence microscopy. RESULTS: Mycoplasma genitalium was shown to adhere to the head, midpiece and tail of the spermatozoa. The spermatozoa became immotile when many M.genitalium were attached. However, the motile spermatozoa were demonstrated to carry M.genitalium and in this case the mycoplasmas were seen to attach mostly to the midpiece or neck region. Occasionally, M.genitalium was seen at the head but not at the tail. By X-ray microscopy, it was possible to observe the diffentiated structure of M.genitalium, and the attachment seemed to be mediated by the tip. CONCLUSIONS: Mycoplasma genitalium can bind to human spermatozoa and thus could be carried by motile sperm. This ability may be important in the process of causing female genital diseases and infertility.     

Mycoplasma pneumoniae induces host-dependent pulmonary inflammation and airway obstruction in mice

Respiratory tract infections result in wheezing in a subset of patients. Mycoplasma pneumoniae is a common etiologic agent of acute respiratory infection in children and adults that has been associated with wheezing in 20-40% of individuals. The current study was undertaken to elucidate the host-dependent pulmonary and immunologic response to M. pneumoniae respiratory infection by studying mice with different immunogenetic backgrounds (BALB/c mice versus C57BL/6 mice). After M. pneumoniae infection, only BALB/c mice developed significant airway obstruction (AO) compared with controls. M. pneumoniae-infected BALB/c mice manifested significantly elevated airway hyperresponsiveness (AHR) compared with C57BL/6 mice 4 and 7 d after inoculation as well as BALB/c control mice. Compared with C57BL/6 mice, BALB/c mice developed worse pulmonary inflammation, including greater peribronchial infiltrates. Infected BALB/c mice had significantly higher concentrations of tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-1beta, IL-6, IL-12, KC (functional IL-8), and macrophage inflammatory protein 1alpha in the bronchoalveolar lavage fluid compared with infected C57BL/6 mice. No differences in IL-2, IL-4, IL-5, IL-10, and granulocyte/macrophage colony-stimulating factor concentrations were found. The mice in this study exhibited host-dependent infection-related AO and AHR associated with chemokine and T-helper type (Th)1 pulmonary host response and not Th2 response after M. pneumoniae infection.     

Cross-hybridization between the cytadhesin genes of Mycoplasma pneumoniae and Mycoplasma genitalium and genomic DNA of Mycoplasma gallisepticum

Immunological cross-reactivity was observed between the cytadhesin proteins of Mycoplasma pneumoniae and Mycoplasma genitalium and a 155 kDa protein of Mycoplasma gallisepticum. Furthermore, the cytadhesin genes of M. pneumoniae and M. genitalium were used to demonstrate homology with M. gallisepticum genomic DNA under low stringency conditions suggesting that a family of adhesin-related genes exists among these pathogenic mycoplasmas.     

Absence of Mycoplasma pneumoniae cytadsorption protein P1 in Mycoplasma genitalium and Mycoplasma gallisepticum

Polyclonal and monoclonal antibodies to Mycoplasma pneumoniae protein P1 were nonreactive with whole-cell or soluble preparations of M. genitalium and M. gallisepticum. However, radioimmunoprecipitation performed with hyperimmune rabbit sera raised against each mycoplasma species indicated antigenic cross-reactivity between M. pneumoniae and M. genitalium.     

Expression and functional identification of the hypothetical adhesin P32 from Mycoplasma genitalium

Mycoplasma genitalium is the main causative agent for non-gonococcal and non-chlamydial urethritis. P32 is the putative surface-exposed membrane protein of M. genitalium and it has substaintial identity in amino acid sequence with adhesin protein P30 from M. pnewnoniae. Since M. pneumoniae mutants lacking P30 protein is defective in cytadherence, P32 protein has been proposed to be an essential adhesin implicated in the adherence of M. genitalium to host cells. The prokaryotic expression vector pET-30 ( )/p32 was constructed in the present study, and the recombinant protein was expressed in E. coli and purified under denaturing condition. As demonstrated by the immunoblotting analysis, the recombinant protein could react with rabbit antisera against M. genitalium, and adherence inhibition assays were petformed with antisera against this recombinant protein. It was demonstrated that P32 protein apperared to be an adhesion protein of M. genitalium, thus providing the experimental basis for better understanding of the pathogenesis of M. genitalium infection and for the development of the related vaccines against the infection.     

Morphology of human Fallopian tubes after infection with Mycoplasma genitalium and Mycoplasma hominis--in vitro organ culture study

BACKGROUND: Female infertility can be caused by scarring and occlusion of the Fallopian tubes. Sexually transmitted bacteria can damage the delicate epithelial layer of human Fallopian tubes (HFT). Genital mycoplasmas are associated with human reproductive failure. Yet, there is not enough evidence that mycoplasmas can cause tubal factor infertility. We analysed the effects of infections with Mycoplasma hominis and Mycoplasma genitalium on the HFT epithelium and compared them with the effects of infections with genital pathogens: Chlamydia trachomatis and Neisseria gonorrhoeae. METHODS: We used an in vitro model in which pieces of normal HFT were infected with different bacteria, and the outcome of the infections was analysed by scanning electron microscopy (SEM) and confocal microscopy. RESULTS: The presence of M. hominis did not cause any morphological changes of the epithelium of HFT. Noticeable changes in the morphology of the ciliated cells were observed in M. genitalium-infected tissue. Five days post-infection, the cilia were abnormally swollen and some of the ciliated cells fell off the epithelium. These effects could be inhibited by pre-incubation of M. genitalium with antibody directed against the C-terminal part of the adhesion protein MgPa before infection of HFT organ culture. CONCLUSION: We have shown that the presence of M. genitalium, but not M. hominis, in the HFT organ culture affected the epithelium and resulted in cilia damage. The effect of infection with M. genitalium on the HFT was, however, very moderate when compared with the extensive damage of the epithelium caused by N. gonorrhoeae or C. trachomatis.     

Mycoplasma hominis in pregnancy

One hundred and seventyone antenatal patients were examined for the presence of ;large colony' mycoplasmas in the vagina, and for complement-fixing antibody to Mycoplasma hominis. In 25 patients the findings before and after delivery were compared. In patients from whom M. hominis was grown, antibody was twice as common after delivery, and the development of antibody was sometimes associated with pyrexia and signs of genital tract infection.     

ELISA法检测江门地区性病门诊患者生殖支原体抗原

目的应用酶联免疫吸附试验（ELISA）检测性病门诊患者泌尿生殖道生殖支原体（Mg）抗原，探讨江门地区性传播疾病（STD）门诊患者Mg感染状况。方法采用Mg多克隆抗体，标记酶结合物，建立检测Mg抗原的双抗体夹心ELISA法，对性病门诊患者泌尿生殖道分泌物进行MIg抗原检测。结果双抗体夹心ELISA法可检测到5μg／ml蛋白浓度，除肺炎支原体外与其他泌尿生殖道常见支原体和细菌无交叉反应；共检测了174例标本，Mg抗原阳性33例，阳性率为18．97％，其中男性和女性阳性率分别为19．88％和7．69％；男性患者中，非淋菌性尿道炎（NGU）、淋病、前列腺炎患者的Mg抗原阳性率分别为13．43％、62．5％和4．84％。结论STD门诊患者存在Mg感染，应用双抗体夹心ELISA法检测Mg抗原具有一定的灵敏度，且具有快速、简便的特点，适合临床筛查Mg抗原。     

Mycoplasma hominis in female genital tract

In dependence on health of women the female genital tract is colonised by different microorganisms. Mycoplasma hominis was the first mycoplasma of human origin to be isolated. M. hominis, a common inhabitant of the vagina of healthy women, becomes pathogenic once it invades the internal genital organs. M. hominis is associated with bacterial vaginosis but it is still unclear whether the organism really contributes to a pathological process in which so many different bacteria are involved. The aim of this article is to summarize known information about these microorganisms.     

Elevated cytokine and chemokine levels and prolonged pulmonary airflow resistance in a murine Mycoplasma pneumoniae pneumonia model: a microbiologic, histologic, immunologic, and respiratory plethysmographic profile

Because Mycoplasma pneumoniae is hypothesized to play an important role in reactive airway disease/asthma, a comprehensive murine model of M. pneumoniae lower respiratory infection was established. BALB/c mice were intranasally inoculated once with M. pneumoniae and sacrificed at 0 to 42 days postinoculation. All mice became infected and developed histologic evidence of acute pulmonary inflammation, which cleared by 28 days postinoculation. By contrast, M. pneumoniae persisted in the respiratory tract for the entire 42 days studied. Tumor necrosis factor alpha, gamma interferon, interleukin-6 (IL-6), KC (functional IL-8), MIP-1alpha, and MCP-1/JE concentrations were significantly elevated in bronchoalveolar lavage samples, whereas IL-4 and IL-10 concentrations were not significantly elevated. Pulmonary airflow resistance, as measured by plethysmography, was detected 1 day postinoculation and persisted even after pulmonary inflammation had resolved at day 28. Serum anti-M. pneumoniae immunoglobulin G titers were positive in all mice by 35 days. This mouse model provides a means to investigate the immunopathogenesis of M. pneumoniae infection and its possible role in reactive airway disease/asthma.