Chlamydia Trachomatis Mouse Pneumonitis Research Articles

Immunity to Chlamydia trachomatis Mouse Pneumonitis Induced by Vaccination with Live Organisms Correlates with Early Granulocyte-Macrophage Colony-Stimulating Factor and Interleukin-12 Production and with Dendritic Cell-Like Maturation

Immunity to Chlamydia trachomatis Mouse Pneumonitis Induced by Vaccination with Live Organisms Correlates with Early Granulocyte-Macrophage Colony-Stimulating Factor and Interleukin-12 Production and with Dendritic Cell-Like Maturation

Different roles are played by alpha beta and gamma delta T cells in acquired immunity to Chlamydia trachomatis pulmonary infection.

Using gene knockout and wild-type C57BL/6 mice, we examined the role of alpha beta and gamma delta T cells in the resolution of Chlamydia trachomatis mouse pneumonitis (MoPn) biovar pulmonary infection. The results show that alpha beta T-cell-deficient (alpha-/-) mice, when compared with wild-type control mice, have dramatically increased mortality rate and greater in vivo growth of MoPn. The alpha beta T-cell-deficient mice were as susceptible to MoPn infection as T- and B-lymphocyte-deficient (RAG-1-/-) mice. Moreover, both alpha beta T-cell-deficient and RAG-1 mutant mice failed to mount delayed-type hypersensitivity (DTH) responses to organism-specific challenge and showed undetectable interferon-gamma (IFN-gamma) production by spleen cells upon in vitro organism-specific restimulation. In contrast, gamma delta T-cell-deficient mice exhibited intact DTH responses and their mortality rate and in vivo chlamydial growth were comparable to those in wild-type controls. More interestingly, gamma delta T-cell-deficient mice showed significantly higher levels of IFN-gamma production than did wild-type mice. The data indicate that the alpha beta T cell is the major T-cell population for acquired immunity to chlamydial infection and that gamma delta T cells may play an ancillary role in regulating the magnitude of alpha beta T-cell responses.

DNA vaccination with the major outer-membrane protein gene induces acquired immunity to Chlamydia trachomatis (mouse pneumonitis) infection.

The efficacy of DNA vaccination for prevention of Chlamydia trachomatis infection was studied using the murine model of pneumonia induced by the mouse pneumonitis (MoPn) isolate of C. trachomatis. Intramuscular DNA immunization with two chlamydial genes, one that encodes the major outer-membrane protein (MOMP) and one that encodes a cytoplasmic enzyme (cytosine triphosphate [CTP] synthetase) were tested. The MOMP DNA vaccine but not the CTP synthetase DNA vaccine generated significant delayed-type hypersensitivity and serum antibodies to MoPn elementary bodies and reduced the peak growth of MoPn by >100-fold following lung challenge infection. MOMP DNA immunization suggests a new approach to vaccine development for prevention of human chlamydial infection.

Immunization with an acellular vaccine consisting of the outer membrane complex of Chlamydia trachomatis induces protection against a genital challenge

The ability to induce protection against a genital challenge was studied in BALB/c female mice with three Chlamydia trachomatis mouse pneumonitis (MoPn) major outer membrane protein (MOMP) preparations as well as an acellular vaccine consisting of the chlamydial outer membrane complex (COMC). The MOMP preparations were extracted with three different types of detergents, sodium dodecyl sulfate (SDS), n-octyl-beta-D-glucopyranoside (OGP), and Zwittergent 3-14 (Z3-14). A positive immunization control consisted of mice inoculated intranasally with 10(4) C. trachomatis MoPn inclusion-forming units (IFU). Mice inoculated with ovalbumin served as a negative control. Furthermore, a sham-immunized, nonchallenged group was included as a fertility control. Two weeks after the last immunization, the mice were challenged in the left ovarian bursa with 10(5) C. trachomatis MoPn IFU. Vaginal swabs were collected for culture, vaginal and serum samples were assayed for chlamydial-specific antibodies, and splenocytes were collected to determine the lymphoproliferative response. At 42 days after the challenge, the mice were mated with proven male breeder mice. Animals that were considered to be pregnant (as determined by weight) were killed, and the embryos were counted. A significant humoral and cell-mediated immune response was observed in all the groups of mice inoculated with chlamydial antigens. Antibodies to variable domain (VD)1 of the MOMP were detected in serum samples from all the immunized groups. However, antibodies to VD3 and VD4 were detected only in the groups immunized with the Z3-14-MOMP and the COMC. Mice immunized with COMC developed significant immunoglobulin A chlamydia-specific antibodies in the vagina, while mice immunized with the detergent-extracted MOMPs had low antibody titers. Following the intrabursal challenge, a significant decrease in the intensity and duration of vaginal shedding was noted in the mice immunized with COMC and a moderate decrease was noted in the group immunized with OGP-MOMP. No protection against the infection was noted in the groups of animals immunized with SDS- and Z3-14-MOMP. Furthermore, of the mice immunized with the COMC preparation, only 25% (4 of 20) shed C. trachomatis, as determined by vaginal culture, while 83% (40 of 48) of the control mice inoculated with ovalbumin were culture positive (P < 0.05). In addition, after mating, the mice inoculated with COMC were found to have fertility rates comparable to those of the control sham-immunized, nonchallenged animals (70% [14 of 20] versus 81% [17 of 21], respectively [P > 0.05]), and there were no significant differences between the average number of embryos per mouse in the two groups (5.1 versus 5.9, respectively [P > 0.05]). In contrast, mice immunized with the purified MOMP preparations were not protected against infertility. In summary, a preparation of the COMC protected mice against infection and infertility, supporting the feasibility of the development of an acellular vaccine against C. trachomatis infections.

Monoclonal immunoglobulin A antibody to the major outer membrane protein of the Chalamydia trachomatis mouse pneumonitis biovar protects mice against a chlamydial genital challenge

In order to analyze the protective role that IgA may play in a chlamydial infection two IgA monoclonal antibodies (mAb), MoPn 4-2 and MoPn 13-2, were raised against the major outer membrane protein (MOMP) of the Chlamydia trachomatis mouse pneumonitis (MoPn) biovar. mAb MoPn 4-2 was found to be serovar specific while mAb MoPn 13-2 was species specific. mAb MoPn 4-2 recognized a surface exposed conformational epitope as shown by its ability to bind to native EBs and nonreduced MOMP while failing to bind to heat and trypsin treated EBs, to reduced MOMP and to synthetic MOMP peptides. In contrast, mAb MoPn 13-2 recognized a nonconformational epitope since it was able to bind treated EBs, to reduced MOMP and to the synthetic peptide MTTWNPTISGSGI located in variable domain 4 of the MOMP. Both mAbs agglutinated intact EBs and had in vitro neutralizing activity. However, mAb MoPn 4-2 had a 20-fold higher in vitro neutralizing ability when compared to mAb MoPn 13-2 (50% neutralization at 5 μg ml −1 vs 100 μg ml −1). In an in vitro-in vivo infectivity assay, mAb MoPn 4-2 protected mice against infertility when C. trachomatis MoPn elementary bodies were preincubated with the mAb before inoculation. In addition, passive transfer of mAb MoPn 4-2 resulted in significant protection as measured by a decrease in the number of mice infected, and in the intensity and duration of vaginal shedding. These results support previous findings suggesting that IgA antibodies can play a role in protection against a chlamydial infection, and further encourage work to develop vaccination strategies that elicit mucosal immunity.

Intranasal immunization induces long-term protection in mice against a Chlamydia trachomatis genital challenge.

In an attempt to confer long-term protective immunity, BALB/c female mice were immunized intranasally with 10(4) inclusion-forming units (IFU) of the Chlamydia trachomatis mouse pneumonitis biovar (MoPn). Animals were subsequently challenged in the ovarian bursa with 10(5) C. trachomatis MoPn IFU at 60, 120, or 180 days post-intranasal immunization. Two control groups were included in the study. One control was sham immunized and mock challenged, and another group was sham immunized and challenged with 10(5) C. trachomatis MoPn IFU. Vaginal cultures were collected at regular intervals following the intrabursal challenge. In comparison with the sham-immunized mice, the animals that were intranasally immunized with C. trachomatis had significant protection, as shown by a reduction in the number of animals that had positive vaginal cultures and by a decrease in the intensity and length of the shedding. Furthermore, histopathological characterization of the genital tract following challenge, in the three groups of mice, showed a minimal inflammatory infiltrate in the C. trachomatis-immunized animals, when compared with the sham-immunized control group. Subsequently, the three groups of female mice that were challenged at 60, 120 and 180 days postimmunization were mated at 6 weeks following the challenge. Overall, in the mice intranasally immunized with C. trachomatis the fertility rates and the number of embryos were similar to those in the sham-immunized and mock-challenged group. In contrast, there was a significant increase in infertility in the groups of mice that were sham immunized and C. trachomatis challenged. In conclusion, intranasal immunization with C. trachomatis induces long-term protection against a genital challenge as shown by a decrease in the infection and infertility rates when compared with sham-immunized animals. Thus, this model may help to characterize the parameters of the immune response that are important in maintaining long-term protection and may aid in identifying the antigenic determinants involved in eliciting protection.

Genetically determined differences in IL-10 and IFN-gamma responses correlate with clearance of Chlamydia trachomatis mouse pneumonitis infection.

Cytokine patterns elicited by infection are critical in the regulation of the adaptive immune response and in the resolution of infection. Using a murine model of pneumonia induced by intranasal inoculation with the Chlamydia trachomatis mouse pneumonitis (MoPn) biovar, we found that the patterns of immune responses and cytokine production by spleen cells were correlated with quantitative growth of MoPn in the lungs of C57BL/6 and BALB/c mice. Specifically, BALB/c (H-2d) mice had a significantly higher mortality rate and a slower clearance of the organism from the lungs than did C57BL/6 mice (H-2b). BALB/c mice exhibited higher IL-10 production, higher IgG1 Ab responses, and less IFN-gamma production than C57BL/6 mice, which showed higher IFN-gamma production, stronger delayed-type hypersensitivity (DTH) responses, and significantly less IL-10 production. In vivo neutralization of IL-10 in BALB/c mice with an anti-IL-10 mAB during MoPn infection significantly increased the DTH response and enhanced clearance of MoPn. These findings support the hypothesis that excessive IL-10 production in BALB/c mice inhibits Th1-like responses, including IFN-gamma expression and the DTH response following chlamydial infection and consequently delay infection resolution.

Intravaginal inoculation of mice with the Chlamydia trachomatis mouse pneumonitis biovar results in infertility.

In an attempt to establish a model of chlamydial ascending salpingitis and infertility, three inbred strains of mice, C3H/HeN (H-2k), C57BL/6N (H-2b), and BALB/cAnN (H-2d), were inoculated intravaginally with 3 x 10(7) inclusion-forming units of the Chlamydia trachomatis mouse pneumonitis biovar. Mice mated 6 weeks following inoculation were found to have a significant decrease in fertility rate compared with the control groups, as shown by a reduction in the number of pregnant mice and a decrease in the number of embryos.

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.

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.

Analysis of the immune response in mice following intrauterine infection with the Chlamydia trachomatis mouse pneumonitis biovar.

A Swiss Webster white mouse model of salpingitis was used to characterize the immune response following an intrauterine infection with the Chlamydia trachomatis mouse pneumonitis biovar. Western blot (immunoblot) analyses of the serum samples showed that the immunodominant bands corresponded to molecular masses of 72, 60, 42, and 28 kDa and to the lipopolysaccharide. Antibodies to the 60-kDa heat shock protein and to the 60-kDa cysteine-rich protein were detected at 2 and 3 weeks postinfection, respectively. Neutralization was observed in an in vitro assay with serum samples as early as the 3rd day postinfection and remained high for the 7 weeks of observation. The mice were mated in the 7th week following infection. Of the infected experimental mice, 71.4% were found to be either unilaterally or bilaterally infertile, whereas only 27.4% of the noninfected control mice were found to be infertile.

Sequence of the gene encoding the major outer membrane protein of the mouse pneumonitis biovar of Chlamydia trachomatis

The gene encoding the major outer membrane protein of the Chlamydia trachomatis mouse pneumonitis biovar was sequenced and the amino acid sequence deduced. The primary structure of this protein is similar to that of the lympho-granuloma venereum and trachoma biovars in that it consists of four variable domains interspersed with five constant domains. This protein may be an ideal candidate for a vaccine in chlamydia-infected mouse experimental models.

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.

Determination of the number oftufgenes inChlamydia trachomatisandNeisseria gonorrhoeae

Restriction endonuclease fragments of DNA from Neisseria gonorrhoeae and Chlamydia trachomatis (mouse pneumonitis biovar) were hybridized to probes from the N-terminal and C-terminal portions of the Escherichia coli tufA gene. In common with other Gram-negative bacteria, the genome of N. gonorrhoeae was found to contain two homologous sequences (presumptive tuf genes). The C. trachomatis genome contained a single tuf sequence.

Determination of the number of tuf genes in Chlamydia trachomatis and Neisseria gonorrhoeae

Restriction endonuclease fragments of DNA from Neisseria gonorrhoeae and Chlamydia trachomatis (mouse pneumonitis biovar) were hybridized to probes from the N-terminal and C-terminal portions of the Escherichia coli tufA gene. In common with other Gram-negative bacteria, the genome of N. gonorrhoeae was found to contain two homologous sequences (presumptive tuf genes). The C. trachomatis genome contained a single tuf sequence.

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.

Activity of Oral Amoxicillin, Ampicillin, and Oxytetracycline against Infection with Chlamydia trachomatis in Mice

The effects in mice of oral treatment with amoxicillin, ampicillin, and oxytetracycline against an otherwise lethal intranasal infection with Chlamydia trachomatis (mouse pneumonitis) were studied. When treatment was started 30 min after infection and continued once daily thereafter for a total of seven treatments, the mean protective doses of amoxicillin, ampicillin, and oxytetracycline were 9.5, greater than 50, and 31.3 mg/kg, respectively. If 14 oral treatments were given, these values were 1.6 mg/kg for amoxicillin, 12.7 mg/kg for ampicillin, and 12.3 mg/kg for oxytetracycline.