Vaccination with the recombinant major outer membrane protein elicits long-term protection in mice against vaginal shedding and infertility following a Chlamydia muridarum genital challenge.

Implementation of a vaccine is likely the best approach to curtail Chlamydia trachomatis infections. The aim of this study was to determine the ability of a vaccine formulated with the recombinant major outer membrane protein (MOMP) and Th1 and Th2 adjuvants, delivered by combinations of systemic and mucosal routes, to elicit long-term protection in mice against a genital challenge with Chlamydia muridarum. As a negative control, mice were vaccinated with the recombinant Neisseria gonorrhoeae porinB, and the positive control group was immunized with C. muridarum live elementary bodies (EB). The four vaccines formulated with MOMP, as determined by the titers of IgG and neutralizing antibodies in serum, proliferative responses of T-cells stimulated with EB and levels of IFN-γ in the supernatants, elicited robust humoral and cellular immune responses over a 6-month period. Groups of mice were challenged genitally at 60, 120, or 180 days postimmunization. Based on the number of mice with positive vaginal cultures, number of positive cultures, length of time of shedding, and number of inclusion forming units recovered, MOMP vaccinated groups were significantly protected. To assess fertility, when the vaginal cultures became negative, female mice were caged with male mice and the outcome of the pregnancy evaluated. As determined by the number of pregnant mice and the number of embryos, two of the vaccine formulations protected mice up to 180 days postimmunization. To our knowledge this is the first subunit of Chlamydia vaccine that has elicited in mice significant long-term protection against a genital challenge.

Proteomic identification of immunodominant chlamydial antigens in a mouse model.

Chlamydia trachomatis is the most common bacterial sexually transmitted pathogen in the world. To identify new vaccine candidates a protein microarray was constructed by expressing the open reading frames (ORFs) from Chlamydia mouse pneumonitis (MoPn). C57BL/6, C3H/HeN and BALB/c mice were immunized either intranasally or intravaginally with live MoPn elementary bodies (EB). Two additional groups were immunized by the intramuscular plus subcutaneous routes with UV-treated EB, using CpG and Montanide as adjuvants to favor a Th1 response, or Alum, to elicit a Th2 response. Serum samples collected from the three strains of mice were tested in the microarray. The array included the expression of 909 proteins from the 921 ORFs of the MoPn genome and plasmid. A total of 530 ORFs were recognized by at least one serum sample. Of these, 36 reacted with sera from the three strains of mice immunized with live EB. These antigens included proteins that were previously described as immunogenic such as MOMP and HSP60. In addition, we uncovered new immunogens, including 11 hypothetical proteins. In summary, we have identified new immunodominant chlamydial proteins that can be tested for their ability to induce protection in animal models and subsequently in humans.

Induction of protection in mice against a respiratory challenge by a vaccine formulated with the Chlamydia major outer membrane protein adjuvanted with IC31®.

IC31(®), a novel adjuvant, has been shown to be effective by increasing the levels of IFN-γ in animal models when delivered with several antigens. Here, we tested the ability of IC31(®), to enhance the protective ability of the Chlamydia trachomatis native major outer membrane protein (nMOMP). BALB/c mice were immunized by the intramuscular (i.m.) and subcutaneous (s.c.) routes with nMOMP+IC31(®). Another group of animals was immunized with nMOMP+Alum and as a negative control mice were immunized with ovalbumin (Ova)+IC31(®). Animals immunized with nMOMP+IC31(®) developed high Chlamydia-specific IgG titers. The serum levels of IgG1 were higher than those of the IgG2a. T cells, from the spleens of mice immunized with IC31(®)-adjuvanted nMOMP demonstrated a strong lymphoproliferative reaction to Chlamydia elementary bodies (EB) compared with the groups immunized with nMOMP+Alum or Ova+IC31(®). A similar comparison between these groups of mice revealed that the levels of IFN-γ in the supernatants from stimulated T-cells were significantly higher in animals immunized with nMOMP+IC31(®). Following an intranasal challenge with C. trachomatis, the mice immunized with IC31(®)-adjuvanted nMOMP showed significant protection. The change in body weight, an indication of the severity of the infection, was significantly less reduced in mice immunized with nMOMP+IC31(®). Furthermore, the weight of the lungs, as well as the pulmonary Chlamydia burden, was significantly lower in the animals immunized with nMOMP+IC31(®) when compared with the groups immunized with nMOMP+Alum or with Ova+IC31(®). In conclusion, these results provide the rationale for further preclinical testing of IC31(®) using other chlamydial antigens, and support the potential evaluation of this adjuvant in human vaccines against Chlamydia.

Identification of immunodominant antigens by probing a whole Chlamydia trachomatis open reading frame proteome microarray using sera from immunized mice.

Chlamydia trachomatis infections can lead to severe chronic complications, including trachoma, ectopic pregnancy, and infertility. The only effective approach to disease control is vaccination. The goal of this work was to identify new potential vaccine candidates through a proteomics approach. We constructed a protein chip array (Antigen Discovery, Inc.) by expressing the open reading frames (ORFs) from C. trachomatis mouse pneumonitis (MoPn) genomic and plasmid DNA and tested it with serum samples from MoPn-immunized mice. Two groups of BALB/c female mice were immunized either intranasally or intravaginally with live elementary bodies (EB). Another two groups were immunized by a combination of the intramuscular and subcutaneous routes with UV-treated EB (UV-EB), using either CpG and Montanide as adjuvants to favor a Th1 response or alum to elicit a Th2 response. Serum samples collected at regular intervals postimmunization were tested in the proteome array. The microarray included the expression products of 909 proteins from a total of 921 ORFs of the Chlamydia MoPn genome and plasmid. A total of 185 immunodominant proteins elicited an early and sustained antibody response in the mice immunized with live EB, and of these, 71 were also recognized by the sera from mice immunized with UV-EB. The reactive antigens included some proteins that were previously described as immunogenic, such as the major outer membrane protein, OmpB, Hsp60, and IncA and proteins from the type III secretion system. In addition, we identified in mice several new immunogens, including 75 hypothetical proteins. In summary, we have identified a new group of immunodominant chlamydial proteins that can be tested for their ability to induce protection.

Induction of protective immunity by vaccination against Chlamydia trachomatis using the major outer membrane protein adjuvanted with CpG oligodeoxynucleotide coupled to the nontoxic B subunit of cholera toxin.

The present study was undertaken to test the efficacy of immunization with the native major outer membrane protein (nMOMP) of Chlamydia trachomatis mouse pneumonitis (MoPn) serovar in combination with a novel immunostimulatory adjuvant consisting of CpG oligodeoxynucleotide (ODN) linked to the nontoxic B subunit of cholera toxin (CTB-CpG) to elicit a protective immune response to C. trachomatis. High levels of Chlamydia-specific IgG antibodies were detected in the sera from BALB/c mice immunized intramuscularly and subcutaneously (i.m.+s.c.) with the nMOMP/CTB-CpG vaccine or with nMOMP adjuvanted with a mixture of CT and CpG ODN (CT+CpG). Further, these immunization schemes gave rise to significant T-cell-mediated Chlamydia-specific immune responses. No Chlamydia-specific humoral or cell-mediated immune responses were detected in the control mice vaccinated with ovalbumin together with either CTB-CpG or CT+CpG. Following an intranasal challenge with C. trachomatis the groups of mice immunized with nMOMP plus CTB-CpG, CT+CpG or live C. trachomatis were found to be protected based on their change in body weight and lung weight as well as number of inclusion forming unit recovered from the lungs, as compared with control groups immunized with ovalbumin plus either adjuvants. Interestingly, IFN-gamma-producing CD4(+), but not CD8(+), T-cells showed a significant correlation with the outcomes of the challenge. In conclusion, nMOMP in combination with the novel adjuvant CTB-CpG elicited a significant antigen-specific antibody and cell-mediated immune responses as well as protection against a pulmonary challenge with C. trachomatis.