Pneumonitis Agent Research Articles

The sequence of the groES and groEL genes from the mouse pneumonitis agent of Chlamydia trachomatis

The co-transcribed structural genes, groES and groEL, of the groE stress response operon from Chlamydia trachomatis mouse pneumonitis (MoPn), were cloned and sequenced. The calculated molecular masses of the encoded heat-shock proteins (Hsp), a small Hsp (GroES) and Hsp60 (GroEL), are 11089 and 58 367 Da, respectively. By comparison with other known chlamydial groES and groEL sequences, there is 89 and 94% nucleotide (nt) identity with C. trachomatis human strains (serovars A and L2), 77 and 82% with C. psittaci strain GPIC, and 75 and 80% with C. pneumoniae isolate AR-39. At the amino-acid level, the MoPn Hsp60 shows a 99% identity with those from C. trachomatis human strains. In a mouse model, MoPn Hsp60 could prove useful in deciphering the pathogenesis of human chlamydial diseases.

Disseminated Trichosporon beigelii infection in patients with malignant diseases: immunohistochemical study and review.

Trichosporon beigelii is a causative agent of opportunistic infection and summer-type hypersensitivity pneumonitis in Japan. However, as the diagnosis of Trichosporon beigelii infection is sometimes difficult, the actual incidence of this disease may be underestimated. Of 203 autopsy patients with malignant disease, seven (7.7%) were diagnosed with disseminated Trichosporon beigelii infection by immunohistochemical investigation of formalin-fixed, paraffin-embedded tissue sections. Including these seven, a total of 43 patients with Trichosporon beigelii infection have been reported in Japan. The majority of them had underlying hematologic malignancies, for which they received cytotoxic chemotherapy resulting in neutropenia. This study indicates that the immunohistochemical method, which can be applied to biopsy specimens, is an excellent tool for specific diagnosis of Trichosporon beigelii infection, which is an emerging fatal mycosis in immunocompromised patients with profound neutropenia.

Gamma interferon levels during Chlamydia trachomatis pneumonia in mice.

Host defense against murine Chlamydia trachomatis (mouse pneumonitis agent [MoPn]) in a murine model was investigated. Gamma interferon (IFN-gamma) was produced in the lungs by both MoPn-susceptible nude athymic (nu/nu) and MoPn-resistant heterozygous (nu/+) mice. In vivo depletion of IFN-gamma in nu/nu mice led to exacerbation of infection. Fluorescence-activated cell sorter analysis disclosed induction of GL3 antibody-positive cells (putatively gamma/delta+ T cells) in nu/nu mouse lung during infection with MoPn. Treatment of nu/nu mice in vivo with antibody to NK cells (anti-asialo GM1 antibody) or to gamma/delta cells (UC7-13D5) did not significantly decrease IFN-gamma production in the lung. However, treatment of severe combined immunodeficiency mice (which lack gamma/delta cells) with antibody to NK cells significantly reduced lung IFN-gamma levels.

Molecular biology of chlamydiae.

Chlamydiae are obligate, intracellular bacteria which cause disease in many species of mammals and birds. There are three species of chlamydiae, Chlamydia trachomatis, Chlamydia pneumoniae and Chlamydia psittaci. Chlamydiae which cause disease in humans are listed in Table 2.1. C. trachomatis is a common human mucosal pathogen which causes three forms of disease; trachoma, genital infection and lymphogranuloma venereum (LGV).The only non-human strain is the murine pathogen, mouse pneumonitis agent. C. trachomatis is divided into 15 serovars, A, B, Ba, C, D, E, F, G, H, I, J, K, L1, L2 and L3. These serovars are grouped into two sub-species; the B complex which comprises serovars B, Ba, D, E, F, G, L1 and L2, and the C complex which comprises serovars A, C, H, I, J, K and L3. C. pneumoniae includes the human pathogens IOL 207 and TWAR. The first isolates came from the eyes of children with trachoma in Iran and Taiwan; more recent isolates have been obtained from patients with respiratory infection. Many people have serum antibodies to C. pneumoniae, indicating that infection iscommon. Most strains of C. psittaci are pathogens of non-primate mammals and birds; only a few of these strains are human pathogens.

Role of L3T4-bearing T-cell populations in experimental murine chlamydial salpingitis

A role for both the cellular and humoral components of the immune response has been established for chlamydial infection. The significance of helper (L3T4) T cells was evaluated by using a Chlamydia trachomatis murine salpingitis model for upper genital tract chlamydial infection. Mouse oviducts were inoculated with C. trachomatis by using the mouse pneumonitis agent (MoPn) or control medium. Mice depleted of L3T4-bearing lymphocytes had significantly higher (P less than 0.05) numbers of organisms recovered at day 7 postinoculation. The rate of hydrosalpinx formation was significantly higher in the mice depleted of L3T4-bearing lymphocytes (27 of 31 [87%] ) than in the infected undepleted group (8 of 16 [50%] ) (P less than 0.01). The geometric mean antichlamydial immunoglobulin G titers at day 54 postinoculation were significantly higher in the L3T4-depleted mice (mean titer, 2,030) than in the undepleted group (mean titer, 776; P less than 0.05). The rate of fertility was lower in the L3T4-depleted group (2 of 31 [6%]) than in the infected, undepleted mice (2 of 16 [13%]), but this difference did not reach statistical significance. In conclusion, the greater persistence of organisms in the oviduct and higher rates of hydrosalpinx formation in mice depleted of L3T4-bearing cells suggests that these cells play a role in the clearing of organisms following infection and thus in reducing the degree of oviduct obstruction and damage.

Resolution of chlamydial genital infection with antigen-specific T-lymphocyte lines

To determine cell-mediated immune mechanisms involved in the resolution of chlamydial genital infection of mice, we utilized an established murine model in which it has been demonstrated that resolution of infection occurs independently of the antibody response. Splenic T lymphocytes were obtained from mice that had previously been immunized with viable elementary bodies of the mouse pneumonitis agent (MoPn), a Chlamydia trachomatis biovar. Antigen-reactive T lymphocytes were maintained and expanded in vitro by frequent restimulation with UV light-inactivated MoPn in the presence of antigen-presenting cells and recombinant interleukin-2 (rIL-2). Flow cytometry indicated that this cell line was at least 92% positive for the pan-specific T-cell marker Thy1.2. Stimulation of the cells in the presence of syngeneic antigen-presenting cells plus MoPn antigen and in the absence of exogenous IL-2 induced the cells to produce IL-2 activity in culture supernatants. Following adoptive transfer, this T-lymphocyte line was effective in inducing resolution of an ongoing MoPn genital infection in congenitally athymic nude mice which otherwise maintain chronic unresolved infections. The line was less efficient in resolving the infection after longer periods in culture. An additional T-lymphocyte line was derived from the spleens of athymic mice that had received the first line and had resolved the infection. These T cells were also capable of inducing resolution of the infection. Lastly, this cell line was treated with specific antibody and complement to delete either CD4+ or CD8+ T lymphocytes in an attempt to enrich for T-cell subpopulations prior to transfer into infected athymic mice. The anti-CD4-treated line was essentially depleted of CD4 cells, while the anti-CD8-treated line was only partially enriched for CD4 cells, with a large proportion of CD8 cells still present. Nude mice that received either of the treated T-cell lines or the parental cell line were capable of resolving the infection, although the line with increased numbers of CD4 cells was more efficient than either the parental line or the CD8 line.

A role in vivo for tumor necrosis factor alpha in host defense against Chlamydia trachomatis

In a mouse model of pneumonia caused by murine Chlamydia trachomatis (mouse pneumonitis agent [MoPn]), tumor necrosis factor alpha (TNF-alpha) antigen and bioactivity were demonstrated in vivo in the lung during MoPn infection in both athymic (nude) and heterozygous (nu/+) mice. Antibody to TNF-alpha that was exogenously given neutralized the TNF-alpha in the lung, significantly accelerated mortality, and caused a borderline increase in MoPn counts in the lung by culture in nu/+ mice. Lipopolysaccharide-induced TNF-alpha activity or injections of recombinant murine TNF-alpha significantly but modestly protected nu/+ mice against MoPn-induced mortality. TNF-alpha is produced in vivo during C. trachomatis infection and plays a role in host defense.

Importance of Serotype-related Antigen in the Induction of Experimental Hypersensitivity Pneumonitis byTrichosporon cutaneumin Rabbits: Correlation between Granulomatous Alveolitis and Cellular and Humoral Immune Responses to Polysaccharide-rich Antigen

We evaluated the immunopathogenic properties of Trichosporon cutaneum, a major etiologic agent of Japanese summer-type hypersensitivity pneumonitis (HP). When the culture filtrate antigen of T. cutaneum was chromatographed on DEAE-cellulose, two peaks of polysaccharide, fractions A and B, were obtained. Fraction B was highly reactive to the specific IgG and IgA antibodies in the serum and bronchoalveolar lavage fluid samples obtained from sensitized rabbits, whereas fraction A was mainly reactive to the IgG antibodies. When the rabbits sensitized by intratracheal injection with the particulate antigen of T. cutaneum were challenged intratracheally by these antigenic fractions, fraction B induced granulomatous alveolitis, but fraction A induced alveolitis rather than granuloma. Lymphocytes of the mediastinal lymph nodes responded prominently to fraction B, but less to fraction A, as assessed by proliferative response. Each of the fractions B obtained from two strains of different serotypes (TIMM 1573, serotype I and TIMM 1318, serotype II) was reactive in enzyme-linked immunosorbent assay to the serum samples from the rabbits sensitized with homologous antigen, but far less to that from the rabbits sensitized with heterologous antigen, that is, the antigenic specificity of fraction B was related to the serotype of T. cutaneum. Fractionation by gel filtration on Sepharose CL-4B revealed that the molecular weight of the antigenic components in fraction B was larger than 1,000,000 and that the components consisted mainly of polysaccharide.(ABSTRACT TRUNCATED AT 250 WORDS)

Humoral immune response to chlamydial genital infection of mice with the agent of mouse pneumonitis.

The objective of this study was to characterize the humoral immune response to chlamydial genital infection of mice with the mouse pneumonitis agent (MoPn). With an enzyme-linked immunoabsorbent assay, immunoglobulin G antibodies to MoPn were first detected in plasma by day 14. Peak plasma antibody concentrations were reached by day 49, and this response did not decline significantly throughout the 300-day monitoring period. Immunoglobulin A against MoPn could first be detected in pooled vaginal washes by day 21 after infection and had reached peak concentrations by day 28, but anti-MoPn immunoglobulin G was not consistently present in secretions. The antibody response in secretions had declined slightly by day 300. Immunoblot analysis revealed that the early phase of the plasma antibody response to MoPn as a result of genital infection was against lipopolysaccharide, the major outer membrane protein, and a 62-kilodalton (kDa) protein. In secretions, early-phase immunoglobulin A antibodies were directed to the major outer membrane protein and lipopolysaccharide. Late reactions to 15-, 22-, and 83-kDa proteins in plasma were noted. Late reactions to the 62-kDa protein in secretions were also noted. The cause of these late responses remains unexplained. When mice were challenged intravaginally with MoPn at 50-day intervals after the primary infection, it was found that mice inoculated on day 100 or after were susceptible to reinfection. Susceptibility could not be related to a decline in the antibody concentration in plasma or secretions or in the antibody response to specific components of MoPn as measured by immunoblot analysis.

Tumor necrosis factor alpha is a cytotoxin induced by murine Chlamydia trachomatis infection.

A mouse model of pneumonia caused by murine Chlamydia trachomatis (mouse pneumonitis agent) was used to demonstrate that whole spleen cells from both nude athymic mice (nu/nu) and heterozygous mice (nu/+) produced tumor necrosis factor alpha in vitro in response to mouse pneumonitis agent antigen. The tumor necrosis factor alpha measured in these supernatants by immunoassay was shown to have bioactivity in a cytotoxic assay in which uninfected target cells were used. This cytotoxicity was distinct from the gamma interferon-related cytotoxicity against C. trachomatis-infected targets that we described previously.

Pneumocystis cariniiPneumonia in an Infant with the Hyperimmunoglobulinemia E Syndrome*

A 4-month-old female presented with "cold" groin abscesses, otitis, and interstitial pneumonia. Open lung biopsy showed extensive Pneumocystis carinii pneumonitis. An immunologic workup revealed exaggerated blood levels of immunoglobulin E (over 2000 IU/ml by 10 months of age) and abnormal polymorphonuclear chemotaxis, both characteristic of the hyperimmunoglobulinemia E (HIE) syndrome. No well-defined lymphocytic defect known to predispose to opportunistic infection was discovered in this patient. We believe that our patient represents the first reported association of P. carinii pneumonia in a patient with HIE syndrome. This finding has implications for the spectrum of immunologic aberrations that may predispose to P. carinii pneumonia and also should raise consideration for P. carinii as a potential etiologic agent of pneumonitis in patients with HIE syndrome.

Gamma interferon-mediated cytotoxicity related to murine Chlamydia trachomatis infection.

After infection with the mouse pneumonitis agent (MoPn; murine Chlamydia trachomatis), heterozygous (nu/+) but not nude athymic (nu/nu) mice produced enhanced amounts of gamma interferon (IFN-gamma) in vitro in response to MoPn antigen that exhibited cytotoxic activity when added to host cells already infected with chlamydiae. Antibody-complement lysis showed the cytotoxic activity to be dependent, at least in part, on L3T4+ T cells for production. The cytotoxic responses were directed primarily against Chlamydia-infected target cells, but a second type of toxicity was demonstrable against uninfected target cells after treatment of the generating cell population with anti-Lyt-2 antibody plus complement at certain time points after infection. This additional nonspecific cytotoxic activity was presumably due to a second factor (factor X) acting in concert with IFN-gamma. Lyt-2+ cells, however, also were shown to play a role in IFN-gamma production and cytotoxicity directed against infected targets at later time points after infection. Neutralization of IFN-gamma in the samples containing cytotoxic activity abrogated the cytotoxicity against both infected and uninfected targets, but cloned murine IFN-gamma exhibited toxicity in a dose-dependent manner only against infected target cells. The data provides evidence that cytotoxicity against infected targets is due to antigen-specific induction of IFN-gamma, but other cytokine activity, most demonstrable after removal of Lyt-2.2+ cells and cytotoxic to uninfected targets, also is present.

Resolution of chlamydial genital infection in B-cell-deficient mice and immunity to reinfection.

The purpose of this investigation was to determine the relative roles of the humoral and cell-mediated immune responses in the resolution of chlamydial genital infection of mice and resistance to reinfection. To this end, female BALB/c mice were rendered B cell deficient by treatment with heterologous anti-immunoglobulin M (IgM) serum from birth. Controls were similarly treated with either normal serum or phosphate-buffered saline. Before inclusion in each experiment, anti-IgM-treated mice were screened for the absence of IgM in serum and for the presence of cell-mediated immune responses. In addition, spleen cells from anti-IgM-treated mice responded to concanavalin A and phytohemagglutinin but not to lipopolysaccharide. By these criteria, mice were designated B cell deficient. B-cell-deficient mice and controls were inoculated intravaginally with a suspension of mouse pneumonitis agent (MoPn), a Chlamydia trachomatis biovar. All B-cell-deficient mice resolved the infection. Additionally, no significant difference was seen in the course of the infection in B-cell-deficient mice when compared with controls. In contrast to control mice, B-cell-deficient mice displayed no detectable antibody responses to MoPn in serum or in genital secretions. However, both B-cell-deficient mice and controls developed delayed-type hypersensitivity and T-cell proliferative responses to MoPn. When challenged 53 days after primary infection, no significant difference was seen in the resistance of B-cell-deficient mice to reinfection when compared with that of the controls. These data indicate that cell-mediated immune mechanisms play an important role in the resolution of and resistance to chlamydial genital infection in this model.

Environmental mycological studies on the causative agent of summer-type hypersensitivity pneumonitis

Environmental mycological studies were carried out in 22 homes of patients with summer-type hypersensitivity pneumonitis and in 195 homes of control subjects. In 10 patients' homes, indoor sampling was performed by open-plate culture, house dust culture, and swab culture (group 1), but in the other 12 patients' homes, sampling was only by house dust culture (group 2). We isolated 302 strains of yeasts from the 22 patients' homes and 962 strains of yeasts from the homes of control subjects. The incidence of yeasts, except genera Trichosporon, was not significantly different between homes of patients and control subjects when homes were assessed by three culture methods. T. cutaneum was isolated from seven of 10 patients' homes in group 1, and their colonizing places were revealed by the swab culture method. In group 2, the cells were isolated from four of 12 patients' homes. No T. cutaneum, however, was isolated from the control subjects' homes. Among the isolated yeasts from patients' homes, 23 strains were reactive to the patients' sera at 1:128 or higher in indirect fluorescent antibody titers; 10 yeasts were T. cutaneum, isolated from 10 homes of 14 patients, but the other 13 yeasts were each a different species isolated from 10 different homes. Furthermore, inhalation challenge with the culture-filtrate antigen prepared from T. cutaneum was performed on the nine patients of six homes in group 1 and the two asymptomatic family members. Of the nine patients, six were positive, one was probable, and two patients were negative. Neither of the two asymptomatic family members responded. These results suggest that T. cutaneum is a major causative agent of summer-type hypersensitivity pneumonitis.

Primary murine Chlamydia trachomatis pneumonia in B-cell-deficient mice.

Mice were rendered deficient in B-cell activity by treatment with anti-mu antibody from birth. These animals were then infected intranasally with murine Chlamydia trachomatis (murine pneumonitis agent [MoPn]). They produced neither local nor systemic antibody to MoPn but had intact delayed-type hypersensitivity to MoPn. Anti-mu-treated mice were not significantly more susceptible to primary invasive infection with MoPn than were control mice, and unrestricted multiplication with MoPn did not occur. The dominant immune response controlling this type of infection is not likely to be antibody.

Role of natural killer cells in infection with the mouse pneumonitis agent (murine Chlamydia trachomatis).

Natural killer (NK) activity is increased in both spleen and lung early in pulmonary infection by murine Chlamydia trachomatis in both susceptible nude and resistant heterozygous (nu/+) mice. Ablation of the rise in NK activity by giving the mice antiasialo GM-1 antibody or stimulation of NK activity by immunomodulators did not affect quantitative tissue counts of the mouse pneumonitis biovar of C. trachomatis or significantly affect survival. Studies are needed to further define the role of NK cells in host defense, immunoregulation, and immunopathology during chlamydial infection.

Role of cell-mediated immunity in chlamydial infection: implications for ocular immunity.

Cellular immunity to Chlamydia trachomatis is not well understood. Studies with the mouse pneumonitis agent in a mouse pneumonia model suggest a strong thymic component to immunity and a role for the macrophage as an effector cell in cell-mediated immunity to C. trachomatis. Experiments with Chlamydia psittaci indicate that cellular cytotoxic mechanisms and biostatic or cytotoxic mechanisms involving cytokines may also play a role in host defense against chlamydiae. Interferon has been shown to inhibit intracellular growth of both C. trachomatis and C. psittaci in vitro. Further studies are needed on the role of T lymphocytes or lymphokines and macrophages in limiting the growth of C. trachomatis in conjunctival epithelial cells (the most appropriate target cells) and in exerting a toxic effect on these cells. The evidence suggests that macrophages are activated during C. trachomatis infection; this activation may lead to the release of monokines (such as interleukin 1) that may be important in the pathogenesis of scarring due to over-stimulation of cellular immunity during ocular infection. Cell-mediated immunity to ocular infection with C. trachomatis may thus be a double-edged sword.

Antibody in host defense against mouse pneumonitis agent (murine Chlamydia trachomatis).

In a murine model of chlamydial pneumonia employing murine Chlamydia trachomatis, immune serum given intranasally delayed death in nude (nu/nu) mice and prevented death in nu/+ mice. Serum-derived immunoglobulin G and A fractions and immune lavage fluid fractions containing immunoglobulins G and A were effective in producing protection when used to opsonize the murine C. trachomatis inoculum. In hyperclean mice (previously made germfree and then colonized with a limited flora nonpathogenic to mice) antibody given intravenously was ineffective. The effects of antibody and nonspecific stimulation of cell-mediated immunity (after previous infection with Histoplasma capsulatum) were additive in increasing host resistance to murine C. trachomatis.

Hypersensitivity pneumonitis in rabbits. Modulation of pulmonary inflammation by long-term aerosol challenge with antigen

Immunized rabbits that were aerosol challenged for 2 to 3 wk with pigeon dropping extract, an etiologic agent of hypersensitivity pneumonitis, developed chronic pulmonary inflammation associated with cell-mediated immunity in bronchoalveolar cells. However, prolonged aerosol challenge for 12 wk resulted in the diminution of pulmonary inflammation (modulation) and the loss of demonstrable cell-mediated immunity. This was probably not due to loss of sensitized lymphocytes that mediated pulmonary inflammation. Furthermore, rabbits undergoing modulation when they were challenged with an unrelated antigen were refractory to the development of pulmonary inflammation for at least 9 wk. Affer this refractory period, animals reimmunized and aerosol challenged with pigeon dropping extract displayed an anamnestic response and produced pulmonary lesions that were strikingly similar to the histopathology of human hypersensitivity pneumonitis.

Cellular immunity to the mouse pneumonitis agent.

During infection with the mouse pneumonitis biovar of Chlamydia trachomatis, heterozygous (nu/+) mice with relatively intact T cell function develop both delayed hypersensitivity to C trachomatis antigen and antigen-specific lymphocyte transformation, whereas athymic nude (nu/nu) mice do not. Nu/nu mice are protected against death from mouse pneumonitis by transfer of immune T cells from nu/+ mice, which are more resistant to C trachomatis. While this enables athymic mice to make antibody to C trachomatis (which does not occur without reconstitution), resistance correlates best with development of antigen-specific lymphocyte transformation in the recipient animals. During infection nu/+ mice develop activated alveolar macrophages (by both morphological and functional criteria) while nu/nu mice do not. Nu/+ mice that have been preinfected with Histoplasma capsulatum to activate cellular immunity become more resistant to C trachomatis than do nu/+ controls. Cell-mediated immunity to C trachomatis pneumonia is T cell dependent and is important in host defense.